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10.3: Applications of Molecular Markers - Biology

10.3: Applications of Molecular Markers - Biology


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Several characteristics of molecular markers make them useful to geneticists. First, because of the way DNA polymorphisms arise and are retained, they are frequent throughout the genome. Second, because they are phenotypically neutral, it is relatively easy to find markers that differ between two individuals. Third, their neutrality also makes it possible to study hundreds of loci without worrying about gene interactions or other influences that make it difficult to infer genotype from phenotype. Lastly, unlike visible traits such as eye color or petal color, the phenotype of a molecular marker can be detected in any tissue or developmental stage, and the same type of assay can be used to score molecular phenotypes at millions of different loci. Thus, the neutrality, high density, high degree of polymorphism, co-dominance, and ease of detection of molecular markers has lead to their wide adoption in many areas of research.

It is worth emphasizing again that DNA polymorphisms are a natural part of most genomes. Geneticists discover these polymorphisms in various ways, including comparison of random DNA sequence fragments from several individuals in a population. Once molecular markers have been identified, they can be used in many ways, including:

DNA fingerprinting

By comparing the allelic genotypes at multiple molecular marker loci, it is possible to determine the likelihood of similarity between two DNA samples. If markers differ, then clearly the DNA is from different sources. If they don’t differ, then one can estimate the unlikelihood of them coming from different sources – eg they are from the same source. For example, a forensic scientist can demonstrate that a blood sample found on a weapon came from a particular suspect. Similarly, that leaves in the back of a suspect's pick-up truck came from a particular tree at a crime scene. DNA fingerprinting is also useful in paternity testing (Figure (PageIndex{1})) and in commercial applications such as verification of species of origin of certain foods and herbal products.

Construction of genetic linkage maps

By calculating the recombination frequency between pairs of molecular markers, a map of each chromosome can be generated for almost any organism (Figure (PageIndex{2})). These maps are calculated using the same mapping techniques described for genes in Chapter 7, however, the high density and ease with which molecular markers can be genotyped makes them more useful than other phenotypes for constructing genetic maps. These maps are useful in further studies, including map-based cloning of protein coding genes that were identified by mutation.

Population studies

As described in Chapter 5, the observed frequency of alleles, including alleles of molecular markers, can be compared to frequencies expected for populations in Hardy-Weinberg equilibrium to determine whether the population is in equilibrium. By monitoring molecular markers, ecologists and wildlife biologists can make inferences about migration, selection, diversity, and other population-level parameters.

Molecular markers can also be used by anthropologists to study migration events in human ancestry. There is a large commercial business available that will genotype people and determine their deep genetic heritage for ~$100. This can be examined through the maternal line via sequencing their mitochondrial genome and through the paternal line via genotyping their Y-chromosome.

For example, about 8% of the men in parts of Asia (about 0.5% of the men in the world) have a Y-chromosomal lineage belonging to Genghis Khan (the haplogroup C3) and his decendents.

Identification of linked traits

It is often possible to correlate, or link, an allele of a molecular marker with a particular disease or other trait of interest. One way to make this correlation is to obtain genomic DNA samples from hundreds of individuals with a particular disease, as well as samples from a control population of healthy individuals. The genotype of each individual is scored at hundreds or thousands of molecular marker loci (e.g. SNPs), to find alleles that are usually present in persons with the disease, but not in healthy subjects. The molecular marker is presumed to be tightly linked to the gene that causes the disease, although this protein-coding gene may itself be as yet unknown. The presence of a particular molecular polymorphism may therefore be used to diagnose a disease, or to advise an individual of susceptibility to a disease.

Molecular markers may also be used in a similar way in agriculture to track desired traits. For example, markers can be identified by screening both the traits and molecular marker genotypes of hundreds of individuals. Markers that are linked to desirable traits can then be used during breeding to select varieties with economically useful combinations of traits, even when the genes underlying the traits are not known.

Quantitative trait locus (QTL) mapping

Molecular markers can be used to identify multiple different regions of chromosomes that contain genes that act together to produce complex traits. This process involves finding combinations of alleles of molecular markers that are correlated with a quantitative phenotype such as body mass, height, or intelligence. QTL mapping is described in more detail in the following section.


The application of molecular biology

Molecular biology methods have tremendous value not only in the investigation of basic scientific questions, but also in application to a wide variety of problems affecting the overall human condition. Disease prevention and treatment, generation of new protein products, and manipulation of plants and animals for desired phenotypic traits are all applications that are routinely addressed by the application of molecular biology methods. Because of the wide applicability of these methods, they are rapidly becoming a pervasive--some would argue invasive--aspect of our technologically based society. The public concerns that address the application of these methods should be addressed by informed public discussion and debate. While scientists can be extremely critical of the quality, interpretation, and significance of experimental results, they have a rather remarkable tendency to be non-judgmental of the relative social merits of many applications of scientific research. It remains a public responsibility to be sufficiently well-informed to critically assess the merits of applied science research and participate in a communal decision-making process regarding the extent to which a new technology will be allowed to affect society.


  1. Restriction Fragment Length Polymorphism (RFLP)
  2. Amplified Fragment Length Polymorphism (AFLP)
  3. Random Amplified Polymorphic DNA (RAPD)
  4. Cleaved Amplified Polymorphic Sequences (CAPS)
  5. Simple Sequence Repeat (SSR) Length Polymorphism
  6. Single Strand Conformational Polymorphism (SSCP)
  7. Heteroduplex Analysis (HA)
  8. Single Nucleotide Polymorphism (SNP)
  9. Expressed Sequence Tags (EST)
  10. Sequence Tagged Sites (STS)

Restriction Fragment Length Polymorphisms (RFLPs):

RFLPs refer to variations found within a species in the length of DNA fragments generated by specific endonuclease. RFLPs are first type of DNA markers developed to distinguish individuals at the DNA level. RFLP technique was developed before the discovery of Polymerase Chain Reaction (PCR).

The advantages, disadvantages and uses of this technique are presented below:

RFLP technique has several advantages. It is a cheaper and simple technique of DNA sequencing. It does not require special instrumentation. The majority of RFLP markers are co-dominant and highly locus specific. These are powerful tools for comparative and synteny mapping.

It is useful in developing other markers such as CAPS and INDEL. Several samples can be screened simultaneously by this technique using different probes. RFLP genotypes for single copy or low copy number genes can be easily scored and interpreted.

Developing sets of RFLP probes and markers is labour intensive. This technique requires large amount of high quality DNA. The multiplex ratio is low, typically one per gel. The genotyping throughput is low. It involves use of radioactive chemicals. RFLP finger prints for multi-gene families are often complex and difficult to score. RFLP probes cannot be shared between laboratories.

They can be used in determining paternity cases. In criminal cases, they can be used in determining source of DNA sample. They can be used to determine the disease status of an individual. They are useful in gene mapping, germplasm characterization and marker assisted selection. They are useful in detection of pathogen in plants even if it is in latent stage.

Amplified Fragment Length Polymorphism (AFLP):

AFLPs are differences in restriction fragment lengths caused by SNPs or INDELs that create or abolish restriction endonuclease recognition sites. AFLP assays are performed by selectively amplifying a pool of restriction fragments using PCR. RFLP technique was originally known as selective restriction fragment amplification.

It provides very high multiplex ratio and genotyping throughput. These are highly reproducible across laboratories. No marker development work is needed however, AFLP primer screening is often necessary to identify optimal primer specificities and combinations.

No special instrumentation is needed for performing AFLP assays however, special instrumentation is needed for co-dominant scoring.

Start-up costs are moderately low. AFLP assays can be performed using very small DNA samples (typically 0.2 to 2.5 pg per individual). The technology can be applied to virtually any organism with minimal initial development.

The maximum polymorphic information content for any bi-allelic marker is 0.5. High quality DNA is needed to ensure complete restriction enzyme digestion. DNA quality may or may not be a weakness depending on the species. Rapid methods for isolating DNA may not produce sufficiently clean template DNA for AFLP analysis.

Proprietary technology is needed to score heterozygotes and ++ homozygotes. Otherwise, AFLPs must be dominantly scored. Dominance may or may not be a weakness depending on the application.

The homology of a restriction fragment cannot be unequivocally ascertained across genotypes or mapping populations. Developing locus specific markers from individual fragments can be difficult and does not seem to be widely done.

The switch to non-radioactive assays has not been rapid. Chemiluminescent AFLP fingerprinting methods have been developed and seem to work well.

The fingerprints produced by fluorescent AFLP assay methods are often difficult to interpret and score and thus do not seem to be widely used. AFLP markers often densely cluster in centromeric regions in species with large genomes, e.g., barley (Hordeum vulgare L.) and sunflower (Helianthus annuus L.).

This technique has been widely used in the construction of genetic maps containing high densities of DNA marker. In plant breeding and genetics, AFLP markers are used in varietal identification, germplasm characterization, gene tagging and marker assisted selection.

Random Amplified Polymorphic DNA (RAPDs):

RAPD refers to polymorphism found within a species in the randomly amplified DNA generated by restriction endonuclease enzyme. RAPDs are PCR based DNA markers. RAPD marker assays are performed using single DNA primer of arbitrary sequence.

RAPD primers are readily available being universal. They provide moderately high genotyping throughput. This technique is simple PCR assay (no blotting and no radioactivity). It does not require special equipment. Only PCR is needed. The start-up cost is low.

RAPD marker assays can be performed using very small DNA samples (5 to 25 ng per sample). RAPD primers are universal and can be commercially purchased. RAPD markers can be easily shared between laboratories. Locus-specific, co-dominant PCR-based markers can be developed from RAPD markers. It provides more polymorphism than RFLPs.

The detection of polymorphism is limited. The maximum polymorphic information content for any bi-allelic marker is 0.5. This technique only detects dominant markers. The reproducibility of RAPD assays across laboratories is often low. The homology of fragments across genotypes cannot be ascertained without mapping. It is not applicable in marker assisted breeding programme.

This technique can be used in various ways such as for varietal identification, DNA fingerprinting, gene tagging and construction of linkage maps. It can also be used to study phylogenetic relationship among species and sub-species and assessment of variability in breeding populations.

Cleaved Amplified Polymorphic Sequences (CAPS):

CAPS polymorphisms are differences in restriction fragment lengths caused by SNPs or INDELs that create or abolish restriction endonuclease recognition sites in PCR amplicons produced by locus-specific oligonucleotide primers.

CAPS assays are performed by digesting locus-specific PCR amplicons with one or more restriction enzymes and separating the digested DNA on agarose or polyacrylamide gels.

CAPS analysis is versatile and can be combined with single strand conformational polymorphim (SSCP), sequence-characterized amplified region (SCAR), or random amplified polymorphic DNA (RAPD) analysis to increase the chance of finding a DNA polymorphism.

Michaels and Amasino (1998) proposed a variant of the CAPS method called dCAPS based on SNPs.

The genotyping throughput is moderately high. It is a simple PCR assay. Markers are developed from the DNA sequences of previously mapped RFLP markers. Most CAPS markers are co- dominant and locus specific. No special equipment is needed to perform manual CAPS marker assays.

CAPS marker assays can be performed using semi-automated methods, e.g., fluorescent assays on a DNA sequencer (e.g., ABI377). Start-up costs are low for manual assay methods. CAPS assays can be performed using very small DNA samples (typically 50 to 100 ng per individual). Most CAPS genotypes are easily scored and interpreted. CAPS markers are easily shared between laboratories.

Typically, a battery of restriction enzymes must be tested to find polymorphisms. Although CAPS markers still nave great utility and should not be over looked, other methods have emerged as tools for screening locus-specific DNA fragments for polymorphisms, e.g., SNP assays. The development of easily scored and interpreted assays may be difficult for some genes, especially those belonging to multi-gene families.

This is straightforward way to develop PCR-based markers from the DNA sequences of previously mapped RFLP markers. It is a simple method that builds on the investment of an RFLP map and eliminates the need for DNA blotting.

Simple Sequence Repeats (SSRs):

Simple sequence repeats (SSRs) or microsatellites are tandemly repeated mono-, di-, tri-, tetra-, penta-, and hexanucleotide motifs. SSR length polymorphisms are caused by differences in the number of repeats. SSR loci are individually amplified by PCR using pairs of oligonucleotide primers specific to unique DNA sequences flanking the SSR sequence.

Jeffreys (1985) showed that some restriction fragment length polymorphisms are caused by VNTRs. The name “mini satellite” was coined because of the similarity of VNTRs to larger satellite DNA repeats.

SSR markers tend to be highly polymorphic. The genotyping throughput is high. This is a simple PCR assay. Many SSR markers are multi-allelic and highly polymorphic. SSR markers can be multiplexed, either functionally by pooling independent PCR products or by true multiplex- PCR. Semi-automated SSR genotyping methods have been developed. Most SSRs are co-dominant and locus specific.

No special equipment is needed for performing SSRs assays however, special equipment is needed for some assay methods, e.g., semi-automated fluorescent assays performed on a DNA sequences. Start-up costs are low for manual assay methods (once the markers are developed). SSR assays can be performed using very small DNA samples (

100 ng per individual). SSR markers are easily shared between laboratories.

The development of SSRs is labor intensive. SSR marker development costs are very high. SSR markers are taxa specific. Start-up costs are high for automated SSR assay methods. Developing PCR multiplexes is difficult and expensive. Some markers may not multiplex.

SSR markers are used for mapping of genes in eukaryotes.

Single Strand Conformational Polymorphisms (SSCPs):

SSCPs refer to DNA polymorphisms produced by differential folding of single-stranded DNA harboring mutations. The conformation of the folded DNA molecule is produced by intra-molecular interactions and is thus a function of the DNA sequence.

SSCP marker assays are performed using heat-denatured DNA on non-denaturing DNA sequencing gels. Special gels (e.g., mutation detection enhancement gels) have been developed to enhance the discovery of single-strand conformational polymorphisms caused by INDELs, SNPs, or SSRs.

It is a simple PCR assay. Many SSCP markers are multi-allelic and highly polymorphic. Most SSCPs are co-dominant and locus specific. No special equipment is needed. Start-up costs are low. SSCP marker assays can be performed using very small DNA samples (typically 10 to 50 ng per individual).

SSCP markers are easily shared between laboratories. SSCP gels can be silver stained (no radioactivity). The complexity of PCR products can be assessed and individual fragments can be isolated and sequenced.

The development of SSCP markers is labor intensive. SSCP marker development costs can be high. SSCP marker analysis cannot be automated.

SSCPs have been widely used in human genetics to screen disease genes for DNA polymorphisms. Although SSCP analysis does not uncover every DNA sequence polymorphism, the methodology is straight forward and a significant number of polymorphisms can be discovered. SSCP analysis can be a powerful tool for assessing the complexity of PCR products.

Heteroduplex Analysis (HA):

It refers to DNA polymorphisms produced by separating homo-duplex from heteroduplex DNA using non-denaturing gel electrophoresis or partially denaturing high performance liquid chromatography.

Single-base mismatches between genotypes produce hetero-duplexes thus, the presence of hetero-duplexes signals the presence of DNA polymorphisms. Heteroduplex analyses can be rapidly and efficiently performed on numerous genotypes before specific alleles are sequenced, thereby greatly reducing sequencing costs in SNP discovery and SNP marker development.

It is a powerful method for SNP discovery. Automated HA can be performed using HPLC. Most heteroduplex markers are co-dominant and locus specific. HA can be performed using very small DNA samples (typically 10 to 50 ng per individual). HA markers are easily shared between laboratories.

Requires special equipment. One protocol may not be sufficient for heteroduplex analyses of different targets via HPLC.

Heteroduplex analysis has been mostly used in human genetics to screen disease genes for DNA polymorphism. In plant breeding, it is used for detection of pathogens which are in latent stage and thus useful in selection of disease free plants. It is also useful in the discovery of single nucleotide polymorphism.

Single Nucleotide Polymorphism (SNP):

The variations which are found at a single nucleotide position are known as single nucleotide polymorphisms or SNP. Such variation results due to substitution, deletion or insertion. This type of polymorphisms has two alleles and also called bialleleic loci. This is the most common class of DNA polymorphism. It is found both in natural lines and after induced mutagenesis. Main features of SNP markers are given below.

1. SNP markers are highly polymorphic and mostly bialleleic.

2. The genotyping throughput is very high.

3. SNP markers are locus specific.

4. Such variation results due to substitution, deletion or insertion.

5. SNP markers are excellent long term investment.

6. SNP markers can be used to pinpoint functional polymorphism.

7. This technique requires small amount of DNA.

SNP markers are useful in gene mapping. SNPs help in detection of mutations at molecular level. SNP markers are useful in positional cloning of a mutant locus. SNP markers are useful in detection of disease causing genes.

Most of the SNPs are bialleleic and less informative than SSRs. Multiplexing is not possible for all loci. Some SNP assay techniques are costly. Development of SNP markers is labour oriented. More (three times) SNPs are required in preparing genetic maps than SSR markers.

SNPs are useful in preparing genetic maps. They have been used in preparing human genetic maps. In plant breeding, SNPs have been used to lesser extent.

Expressed Sequence Tags (EST):

Expressed Sequence Tags (ESTs) are small pieces of DNA and their location and sequence on the chromosome are known. The variations which are found at a single nucleotide position are known. The term Expressed Sequence Tags (ESTs) was first used by Venter and his colleagues in 1991. Main features of EST markers are given below.

1. ESTs are short DNA sequences (200-500 nucleotide long).

2. They are a type of sequence tagged sites (STS).

3. ESTs consist of exons only.

It is a rapid and inexpensive technique of locating a gene. ESTs are useful in discovering new genes related to genetic diseases. They can be used for tissue specific gene expression.

ESTs have lack of prime specificity. It is a time consuming and labour oriented technique. The precision is lesser than other techniques. It is difficult to obtain large (> 6kb) transcripts. Multiplexing is not possible for all loci.

ESTs are commonly used to map genes of known function. They are also used for phylogenetic studies and generating DNA arrays.

Sequence Tagged Sites (STS):

In genomics, a sequence tagged site (STS) is a short DNA sequence that has a single copy in a genome and whose location and base sequence are known. Main features of STS markers are given below.

1. STSs are short DNA sequences (200-500 nucleotide long).

2. STSs occur only once in the genome.

3. STS are detected by PCR in the presence of all other genomic sequences.

4. STSs are derived from cDNAs.

STSs are useful in physical mapping of genes. This technique permits sharing of data across the laboratories. It is a rapid and most specific technique than DNA hybridization techniques. It has high degree of accuracy. It can be automated.

Development of STS is a difficult task. It is time consuming and labour oriented technique. It require high technical skill.

STS is the most powerful physical mapping technique. It can be used to identify any locus on the chromosome. STSs are used as standard markers to find out gene in any region of the genome. It is used for constructing detailed maps of large genomes.


10.3: Applications of Molecular Markers - Biology

Article Summary:

Molecular Markers Types and Applications
Authors: Vivek Sharma and Ram Kunwar

A genetic marker is a gene or known DNA sequence on a chromosome that can be used to identify individuals or species.

Why we need Molecular Markers
There will be no need if identified traits have these three features
&bull Traits were easily score
&bull Individuals were easily classified into few distinct phenotypic classes
&bull Complete corresponding between phenotypic and genotypes


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Professionals working in diagnostic laboratories, including med techs, molecular pathology residents/fellows, clinical pathologists, clinical chemistry fellows, clinical chemists, and toxicologists

  • List of Contributors
  • Preface, Third Edition
  • Chapter 1. Molecular Diagnostics: Past, Present, and Future
    • 1.1. Introduction
    • 1.2. History of Molecular Diagnostics: Inventing the Wheel
    • 1.3. The Post-Polymerase Chain Reaction Revolution
    • 1.4. Molecular Diagnostics in the Post-Genomic Era
    • 1.5. Future Perspectives: What Lies Beyond
    • 1.6. Conclusions
    • 2.1. Introduction
    • 2.2. History
    • 2.3. Authorization
    • 2.4. Definitions
    • 2.5. Variant Descriptions
    • 2.6. Mutalyzer
    • 2.7. Concluding Remarks
    • 3.1. Introduction
    • 3.2. Genetic Screening Methods
    • 3.3. Genetic Scanning Methods
    • 3.4. Conclusions
    • 4.1. History of the Polymerase Chain Reaction
    • 4.2. Principle of Real-Time Polymerase Chain Reaction
    • 4.3. Real-Time Thermal Cyclers
    • 4.4. How Data Are Obtained
    • 4.5. How Data Are Quantified
    • 4.6. Multiplex Quantitative Polymerase Chain Reaction
    • 4.7. Applications of Quantitative Polymerase Chain Reaction and Reverse Transcriptase-Quantitative Polymerase Chain Reaction
    • 4.8. Criteria for Optimizing Quantitative Polymerase Chain Reaction Assays
    • 4.9. Conclusions
    • 5.1. Introduction
    • 5.2. Commercial Sample-to-Answer Assay Systems
    • 5.3. Clinical Applications: Performance for Infectious Pathogen Diagnostics
    • 5.4. Forensic Applications: Performance for Human Identity Testing
    • 5.5. Continuing Evolution of Sample-to-Answer Technologies
    • 6.1. Introduction to Melting Analysis
    • 6.2. Genotyping of Known Variants by High-Resolution Melting
    • 6.3. Variant Scanning by High-Resolution Melting
    • 6.4. Specific Examples of High-Resolution Melting in Clinical Diagnostics
    • 6.5. Other Applications of High-Resolution Melting in Molecular Diagnostics
    • 6.6. Melting Curve Prediction and Assay Design Tools
    • 6.7. Conclusions
    • 7.1. Introduction
    • 7.2. Clinical Applications of DNA Methylation Analysis
    • 7.3. Methods for DNA Methylation Analysis
    • 7.4. Single-Cell DNA Methylation Analysis
    • 7.5. Conclusions
    • 8.1. Introduction
    • 8.2. Commercially Available Analysis Platforms
    • 8.3. Techniques/Systems in Development
    • 8.4. Potential Future Techniques/Systems/Analysis Platforms
    • 8.5. Perspectives for Future Applications and Diagnostics Techniques
    • 8.6. Conclusions
    • 9.1. Introduction
    • 9.2. Advanced Whole-Genome Sequencing
    • 9.3. What Is Needed to Implement This Vision of Genomic Precision Health Care Fully?
    • 9.4. Conclusion
    • 10.1. Introduction
    • 10.2. Padlock and Selector Probes
    • 10.3. Application of Padlock and Molecular Inversion Probes for Genotyping
    • 10.4. Biosensor Approaches Based on Rolling Circle–Amplified Padlock Probes
    • 10.5. Application of Padlock Probes for Infectious Disease Diagnostics
    • 10.6. Targeted Multiplex Copy Number Variation Analysis Using Selector Probes
    • 10.7. High-Throughput Targeted Sequencing Using Selectors and Gap-Fill Padlock Probes
    • 10.8. In Situ Nucleic Acid Detection Using Padlock Probes
    • 10.9. Automation and Miniaturization of Padlock Probe/Rolling Circle Amplification Assays
    • 10.10. Conclusions
    • 11.1. Overview
    • 11.2. Microfluidics for DNA Amplification and Analysis
    • 11.3. Microfluidics for High-Resolution Melting Analysis
    • 11.4. Microfluidics in Cytogenetics
    • 11.5. Microfluidics for Protein Detection and Analysis
    • 11.6. Microfluidic Sample Preparation
    • 11.7. Microfluidics in Cell Sorting
    • 11.8. Future of Microfluidics for Medical Diagnostics
    • 12.1. Introduction
    • 12.2. Binding the Proteome
    • 12.3. Current Affinity-Based Protein Detection Assays
    • 12.4. Proximity-Dependent Nucleic Acid–Based Assays
    • 12.5. Conclusion and Future Perspectives
    • 13.1. Introduction
    • 13.2. Clinical Impact and “Proteomics” Potential
    • 13.3. Strategies for Mass Spectrometry–Based Proteomics: Discovery and Verification
    • 13.4. Bioinformatics
    • 13.5. Examples of Discovery and Verification Proteomics
    • 13.6. Examples of Protein-Based Diagnostics Assays
    • 13.7. Challenges in Clinical Proteomics
    • 13.8. Future Advances and Concluding Remarks
    • 14.1. Introduction
    • 14.2. From Conventional to Molecular Cytogenetics
    • 14.3. Fluorescence In Situ Hybridization
    • 14.4. Basic Technical Elements and Materials
    • 14.5. Types of Fluorescence In Situ Hybridization Probes and Fluorescence In Situ Hybridization Approaches for Metaphase and Interphase Fluorescence In situ Hybridization
    • 14.6. Multicolor Fluorescence In Situ Hybridization Screening Assays
    • 14.7. Multicolor Whole-Metaphase Scanning Techniques
    • 14.8. Multicolor Chromosome Banding Techniques
    • 14.9. Whole-Genome Scanning and Comparative Genomic Hybridization
    • 14.10. Array-Based Techniques (Microarray)
    • 14.11. Conclusions and Future Perspectives
    • Glossary
    • 15.1. A Tumor Presents
    • 15.2. From Microscopic Examination to Molecular Cytogenomics
    • 15.3. The Promise of Liquid Biopsy for Cancer Diagnostics
    • 15.4. Cytogenomic Applications
    • 15.5. Implementation of Cytogenomics in the Clinic
    • 15.6. Bioinformatics and Data Analysis
    • 15.7. Concluding Remarks
    • 16.1. Introduction
    • 16.2. Pharmacogenetics Versus Pharmacogenomics
    • 16.3. History of Pharmacogenomics
    • 16.4. Analytical Methods in Pharmacogenomics
    • 16.5. Pharmacogenomics in Clinical Settings
    • 16.6. Population Differences in Pharmacogenomics
    • 16.7. Complex Phenotypes
    • 16.8. Pharmacogenomics and Regulatory Agencies
    • 16.9. Pharmacogenomics in Drug Development
    • 16.10. Useful Resources in Pharmacogenomics
    • 16.11. New Trends in Pharmacogenomics
    • 16.12. Ethical Implications
    • 16.13. Public Health Pharmacogenomics
    • 16.14. Conclusions and Future Perspectives
    • 17.1. Introduction
    • 17.2. Nature of Genetic Variation
    • 17.3. Nutritional Epidemiology
    • 17.4. Experimental Models
    • 17.5. Defining the Phenotype
    • 17.6. Integrating Complex Data Sets: Data Management, Bioinformatics, and Statistics
    • 17.7. Conclusions
    • 18.1. Introduction
    • 18.2. DNA Microarrays
    • 18.3. New Developments in DNA Microarrays and Genetic Testing
    • 19.1. Introduction
    • 19.2. Next-Generation Sequencing Pipelines
    • 19.3. Molecular Pathway Analysis: Why and What?
    • 19.4. Conclusions
    • 20.1. Introduction
    • 20.2. Historical Overview of Genetic Databases
    • 20.3. Models for Database Management
    • 20.4. Mutation Database Types
    • 20.5. Locus-Specific Databases in Molecular Genetic Testing
    • 20.6. National/Ethnic Mutation Databases: Archiving the Genomic Basis of Human Disorders on a Population Basis
    • 20.7. Database Management Systems for Locus-Specific Databases and National/Ethnic Mutation Databases
    • 20.8. Incentivizing Data Sharing: The Microattribution Approach
    • 20.9. Future Challenges
    • 20.10. Conclusions
    • 21.1. Introduction
    • 21.2. Genetic Markers Commonly Used for Forensic Analysis
    • 21.3. DNA Extraction Methodologies
    • 21.4. DNA Quantitation
    • 21.5. Capillary Electrophoresis and Data Interpretation
    • 21.6. Statistical Calculations
    • 21.7. Next Generation of Forensic DNA Technologies
    • 21.8. Conclusions
    • 22.1. Introduction
    • 22.2. Classification of Mass Fatalities and Diverse Scenarios for Human Remains Retrieval
    • 22.3. Conventional Identification Criteria Routinely Used for Human Identification
    • 22.4. Criteria for the Preservation of Remains
    • 22.5. DNA Polymorphisms Used for Tracing Kinship Between Fragmentary Human Remains and the Relatives Claiming Them
    • 22.6. Challenges Concerning DNA Degradation and Contamination
    • 22.7. Criteria Evolution and Technical Approaches Applied to DNA-Based Victim Identification in Mass Disasters From the Early 1990s to Date
    • 22.8. Description of Analyzed Cases
    • 22.9. From Forensic Genetics to Forensic Genomics: The Change of a Paradigm Driven by Technology
    • 22.10. Future Perspectives
    • 23.1. Introduction
    • 23.2. Assisted Reproductive Technology and Biopsy
    • 23.3. Preimplantation Genetic Diagnosis for Monogenic Disorders
    • 23.4. Preimplantation Genetic Diagnosis for Chromosomal Aberrations
    • 23.5. Emerging Technologies
    • 23.6. Clinical Outcome of Preimplantation Genetic Diagnosis
    • 23.7. Accuracy and Quality Control
    • 23.8. Conclusions and Future Perspectives
    • Web Resources
    • 24.1. Introduction
    • 24.2. Established Prenatal Screening and Diagnosis Practices
    • 24.3. Historical Background of Noninvasive Prenatal Testing
    • 24.4. Origin of Cell-Free Fetal DNA
    • 24.5. Noninvasive Prenatal Testing Methodologies
    • 24.6. Biological and Technical Factors That Affect Noninvasive Prenatal Testing Results
    • 24.7. Noninvasive Prenatal Testing in Clinical Trials
    • 24.8. Noninvasive Prenatal Testing in the Clinical Setting
    • 24.9. Counseling and Ethical Issues
    • 24.10. Future Applications of Noninvasive Prenatal Testing
    • 24.11. Conclusions
    • 25.1. Introduction
    • 25.2. Getting to the Test: Awareness, Access, and Advertising
    • 25.3. Individual Factors Influencing the Utilization of Genetic Testing
    • 25.4. Getting the Genetic Test Results: Personal Impact and Professional Communication
    • 25.5. Family Communication
    • 25.6. Future Challenges: Complexity and Diversity
    • 26.1. Introduction
    • 26.2. Conclusions and Future Perspectives
    • 27.1. Genetics Literacy and the Public Understanding of Genetic Testing
    • 27.2. Obstacles to Genetics Literacy and How These Might Be Overcome
    • 27.3. Conclusions and Suggestions
    • 28.1. Introduction
    • 28.2. Understanding Regulatory and Other Safety Issues Relevant to Biorepositories
    • 28.3. Individuals Involved in Oversight of a Biorepository
    • 28.4. Safety Training/Employee Education in a Biorepository
    • 28.5. Biorepository Safety Areas
    • 28.6. Conclusions
    • 29.1. Introduction
    • 29.2. International Standards
    • 29.3. Accreditation and Certification
    • 29.4. Elements of a Quality Management System
    • 29.5. Quality Control
    • 29.6. Quality Assessment
    • 29.7. Diagnostic Validation
    • 29.8. Quality Improvement
    • 29.9. Conclusions

    Selectable marker genes in transgenic plants: applications, alternatives and biosafety

    Approximately fifty marker genes used for transgenic and transplastomic plant research or crop development have been assessed for efficiency, biosafety, scientific applications and commercialization. Selectable marker genes can be divided into several categories depending on whether they confer positive or negative selection and whether selection is conditional or non-conditional on the presence of external substrates. Positive selectable marker genes are defined as those that promote the growth of transformed tissue whereas negative selectable marker genes result in the death of the transformed tissue. The positive selectable marker genes that are conditional on the use of toxic agents, such as antibiotics, herbicides or drugs were the first to be developed and exploited. More recent developments include positive selectable marker genes that are conditional on non-toxic agents that may be substrates for growth or that induce growth and differentiation of the transformed tissues. Newer strategies include positive selectable marker genes which are not conditional on external substrates but which alter the physiological processes that govern plant development. A valuable companion to the selectable marker genes are the reporter genes, which do not provide a cell with a selective advantage, but which can be used to monitor transgenic events and manually separate transgenic material from non-transformed material. They fall into two categories depending on whether they are conditional or non-conditional on the presence of external substrates. Some reporter genes can be adapted to function as selectable marker genes through the development of novel substrates. Despite the large number of marker genes that exist for plants, only a few marker genes are used for most plant research and crop development. As the production of transgenic plants is labor intensive, expensive and difficult for most species, practical issues govern the choice of selectable marker genes that are used. Many of the genes have specific limitations or have not been sufficiently tested to merit their widespread use. For research, a variety of selection systems are essential as no single selectable marker gene was found to be sufficient for all circumstances. Although, no adverse biosafety effects have been reported for the marker genes that have been adopted for widespread use, biosafety concerns should help direct which markers will be chosen for future crop development. Common sense dictates that marker genes conferring resistance to significant therapeutic antibiotics should not be used. An area of research that is growing rapidly but is still in its infancy is the development of strategies for eliminating selectable marker genes to generate marker-free plants. Among the several technologies described, two have emerged with significant potential. The simplest is the co-transformation of genes of interest with selectable marker genes followed by the segregation of the separate genes through conventional genetics. The more complicated strategy is the use of site-specific recombinases, under the control of inducible promoters, to excise the marker genes and excision machinery from the transgenic plant after selection has been achieved. In this review each of the genes and processes will be examined to assess the alternatives that exist for producing transgenic plants.


    Wheat quality: Wheat breeding and quality testing in Australia

    Larisa Cato , Daniel Mullan , in Breadmaking (Third Edition) , 2020

    8.3.3 Genomic selection

    Genomic selection (GS) refers to an approach to marker-assisted selection where genetic markers (often SNPs) covering the entire genome are used so that all quantitative trait loci (QTL) of interest are in linkage disequilibrium with at least a single marker. All major wheat breeding companies in Australia are using genomic selection approaches and information to enhance their breeding pipelines to varying degrees. The potential application value of a genomic selection approach is high in wheat quality as it is best utilized for complexly inherited traits. Lower environmental influence is also important even though this still exists in quality testing, with sources recorded for trial year, location, and laboratory testing influences for example. The challenge of GS is to increase the accuracy of selection while decreasing cost so the implementation of the technology can be achieved at an early stage in the breeding process.

    Breeding company genetic gain is partly a function of time so bringing forward confident selection of quality performance allows for the reduction in timeframes and increased genetic gains. The improvements in time can be substantial when GS is implemented with many studies/simulations outlining the benefits. Successful GS relies on the development of accurate models which need to be generated by training data that represents the breeding population and targets. It is difficult, however, to document the real value to Australian programs as there are no reported examples of actual genetic gain in breeding programs as the approach is young and data is commercially protected. Integrating major genes as fixed effects into genomic prediction models has been shown to improve such a genomic selection approach for plant morphological and disease resistance traits ( Bernardo, 2014 Zhao et al., 2015 Arruda et al., 2016 ), and has been verified with the Glu-1 loci markers that were associated with dough rheological traits in academic studies.


    Breeding, molecular markers and molecular biology of the olive tree

    Olive (Olea europaea L.) is a typical crop species of the Mediterranean Basin. A number of cultivars were selected and propagated mainly vegetatively over the centuries for their qualitative and quantitative traits. Due to the long juvenile phase of the tree, few breeding programs have been performed. Therefore the most appropriate process is a selection scheme from heterogeneous populations or cultivars varying in oil quantity and quality, harvest regimes, and biotic and abiotic resistance. Molecular marker techniques have been applied recently on olive to relate, identify, distinguish and characterize different cultivars or genotypes and in order to provide information on olive origin and dispersal and to evaluate olive germplasm for traits with agronomical importance. To understand the regulation of biosynthetic pathways of oil and antioxidants on the molecular level, we have isolated a number of genes encoding for key enzymes in fatty acid and antioxidant biosynthesis, modification and triacylglycerol storage. The gene expression during fruit growth and seed development as well as their transient and temporal expression in different tissues is discussed in relation to storage of fatty acids and to provision of signaling molecules important in plant defense mechanisms and reproduction.


    Contents

    In biology, a probe is a single strand of DNA or RNA that is complementary to a nucleotide sequence of interest.

    RNA probes can be designed for any gene or any sequence within a gene for visualization of mRNA, [3] [4] [5] lncRNA [6] [7] [8] and miRNA in tissues and cells. FISH is used by examining the cellular reproduction cycle, specifically interphase of the nuclei for any chromosomal abnormalities. [9] FISH allows the analysis of a large series of archival cases much easier to identify the pinpointed chromosome by creating a probe with an artificial chromosomal foundation that will attract similar chromosomes. [9] The hybridization signals for each probe when a nucleic abnormality is detected. [9] Each probe for the detection of mRNA and lncRNA is composed of

    20-50 oligonucleotide pairs, each pair covering a space of 40–50 bp. The specifics depend on the specific FISH technique used. For miRNA detection, the probes use proprietary chemistry for specific detection of miRNA and cover the entire miRNA sequence.

    Probes are often derived from fragments of DNA that were isolated, purified, and amplified for use in the Human Genome Project. The size of the human genome is so large, compared to the length that could be sequenced directly, that it was necessary to divide the genome into fragments. (In the eventual analysis, these fragments were put into order by digesting a copy of each fragment into still smaller fragments using sequence-specific endonucleases, measuring the size of each small fragment using size-exclusion chromatography, and using that information to determine where the large fragments overlapped one another.) To preserve the fragments with their individual DNA sequences, the fragments were added into a system of continually replicating bacteria populations. Clonal populations of bacteria, each population maintaining a single artificial chromosome, are stored in various laboratories around the world. The artificial chromosomes (BAC) can be grown, extracted, and labeled, in any lab containing a library. Genomic libraries are often named after the institution in which they were developed. An example being the RPCI-11 library, which is named after Roswell Park Comprehensive Cancer Center (formerly known as Roswell Park Cancer Institute) in Buffalo, New York. These fragments are on the order of 100 thousand base-pairs, and are the basis for most FISH probes.

    Preparation and hybridization process – RNA Edit

    Cells, circulating tumor cells (CTCs), or formalin-fixed paraffin-embedded (FFPE) or frozen tissue sections are fixed, then permeabilized to allow target accessibility. FISH has also been successfully done on unfixed cells. [10] A target-specific probe, composed of 20 oligonucleotide pairs, hybridizes to the target RNA(s). Separate but compatible signal amplification systems enable the multiplex assay (up to two targets per assay). Signal amplification is achieved via series of sequential hybridization steps. At the end of the assay the tissue samples are visualized under a fluorescence microscope.

    Preparation and hybridization process – DNA Edit

    First, a probe is constructed. The probe must be large enough to hybridize specifically with its target but not so large as to impede the hybridization process. The probe is tagged directly with fluorophores, with targets for antibodies or with biotin. Tagging can be done in various ways, such as nick translation, or Polymerase chain reaction using tagged nucleotides.

    Then, an interphase or metaphase chromosome preparation is produced. The chromosomes are firmly attached to a substrate, usually glass. Repetitive DNA sequences must be blocked by adding short fragments of DNA to the sample. The probe is then applied to the chromosome DNA and incubated for approximately 12 hours while hybridizing. Several wash steps remove all unhybridized or partially hybridized probes. The results are then visualized and quantified using a microscope that is capable of exciting the dye and recording images.

    If the fluorescent signal is weak, amplification of the signal may be necessary in order to exceed the detection threshold of the microscope. Fluorescent signal strength depends on many factors such as probe labeling efficiency, the type of probe, and the type of dye. Fluorescently tagged antibodies or streptavidin are bound to the dye molecule. These secondary components are selected so that they have a strong signal.

    FISH is a very general technique. The differences between the various FISH techniques are usually due to variations in the sequence and labeling of the probes and how they are used in combination. Probes are divided into two generic categories: cellular and acellular. In fluorescent "in situ" hybridization refers to the cellular placement of the probe

    Probe size is important because longer probes hybridize less specifically than shorter probes, so that short strands of DNA or RNA (often 10–25 nucleotides) which are complementary to a given target sequence are often used to locate a target. The overlap defines the resolution of detectable features. For example, if the goal of an experiment is to detect the breakpoint of a translocation, then the overlap of the probes — the degree to which one DNA sequence is contained in the adjacent probes — defines the minimum window in which the breakpoint may be detected.

    The mixture of probe sequences determines the type of feature the probe can detect. Probes that hybridize along an entire chromosome are used to count the number of a certain chromosome, show translocations, or identify extra-chromosomal fragments of chromatin. This is often called "whole-chromosome painting." If every possible probe is used, every chromosome, (the whole genome) would be marked fluorescently, which would not be particularly useful for determining features of individual sequences. However, it is possible to create a mixture of smaller probes that are specific to a particular region (locus) of DNA these mixtures are used to detect deletion mutations. When combined with a specific color, a locus-specific probe mixture is used to detect very specific translocations. Special locus-specific probe mixtures are often used to count chromosomes, by binding to the centromeric regions of chromosomes, which are distinctive enough to identify each chromosome (with the exception of Chromosome 13, 14, 21, 22.)

    A variety of other techniques uses mixtures of differently colored probes. A range of colors in mixtures of fluorescent dyes can be detected, so each human chromosome can be identified by a characteristic color using whole-chromosome probe mixtures and a variety of ratios of colors. Although there are more chromosomes than easily distinguishable fluorescent dye colors, ratios of probe mixtures can be used to create secondary colors. Similar to comparative genomic hybridization, the probe mixture for the secondary colors is created by mixing the correct ratio of two sets of differently colored probes for the same chromosome. This technique is sometimes called M-FISH.

    The same physics that make a variety of colors possible for M-FISH can be used for the detection of translocations. That is, colors that are adjacent appear to overlap a secondary color is observed. Some assays are designed so that the secondary color will be present or absent in cases of interest. An example is the detection of BCR/ABL translocations, where the secondary color indicates disease. This variation is often called double-fusion FISH or D-FISH. In the opposite situation—where the absence of the secondary color is pathological—is illustrated by an assay used to investigate translocations where only one of the breakpoints is known or constant. Locus-specific probes are made for one side of the breakpoint and the other intact chromosome. In normal cells, the secondary color is observed, but only the primary colors are observed when the translocation occurs. This technique is sometimes called "break-apart FISH".

    Single-molecule RNA FISH Edit

    Single-molecule RNA FISH, also known as Stellaris® RNA FISH, [11] is a method of detecting and quantifying mRNA and other long RNA molecules in a thin layer of tissue sample. Targets can be reliably imaged through the application of multiple short singly labeled oligonucleotide probes. [12] The binding of up to 48 fluorescent labeled oligos to a single molecule of mRNA provides sufficient fluorescence to accurately detect and localize each target mRNA in a wide-field fluorescent microscopy image. Probes not binding to the intended sequence do not achieve sufficient localized fluorescence to be distinguished from background. [13]

    Single-molecule RNA FISH assays can be performed in simplex or multiplex, and can be used as a follow-up experiment to quantitative PCR, or imaged simultaneously with a fluorescent antibody assay. The technology has potential applications in cancer diagnosis, [14] neuroscience, gene expression analysis, [15] and companion diagnostics.

    Fiber FISH Edit

    In an alternative technique to interphase or metaphase preparations, fiber FISH, interphase chromosomes are attached to a slide in such a way that they are stretched out in a straight line, rather than being tightly coiled, as in conventional FISH, or adopting a chromosome territory conformation, as in interphase FISH. This is accomplished by applying mechanical shear along the length of the slide, either to cells that have been fixed to the slide and then lysed, or to a solution of purified DNA. A technique known as chromosome combing is increasingly used for this purpose. The extended conformation of the chromosomes allows dramatically higher resolution – even down to a few kilobases. The preparation of fiber FISH samples, although conceptually simple, is a rather skilled art, and only specialized laboratories use the technique routinely. [16]

    Q-FISH Edit

    Q-FISH combines FISH with PNAs and computer software to quantify fluorescence intensity. This technique is used routinely in telomere length research.

    Flow-FISH Edit

    Flow-FISH uses flow cytometry to perform FISH automatically using per-cell fluorescence measurements.

    MA-FISH Edit

    Microfluidics-assisted FISH (MA-FISH) uses a microfluidic flow to increase DNA hybridization efficiency, decreasing expensive FISH probe consumption and reduce the hybridization time. MA-FISH is applied for detecting the HER2 gene in breast cancer tissues. [17]

    MAR-FISH Edit

    Microautoradiography FISH is a technique to combine radio-labeled substrates with conventional FISH to detect phylogenetic groups and metabolic activities simultaneously. [18]

    Hybrid Fusion-FISH Edit

    Hybrid Fusion FISH (HF-FISH) uses primary additive excitation/emission combination of fluorophores to generate additional spectra through a labeling process known as dynamic optical transmission (DOT). Three primary fluorophores are able to generate a total of 7 readily detectable emission spectra as a result of combinatorial labeling using DOT. Hybrid Fusion FISH enables highly multiplexed FISH applications that are targeted within clinical oncology panels. The technology offers faster scoring with efficient probesets that can be readily detected with traditional fluorescent microscopes.

    Often parents of children with a developmental disability want to know more about their child's conditions before choosing to have another child. These concerns can be addressed by analysis of the parents' and child's DNA. In cases where the child's developmental disability is not understood, the cause of it can potentially be determined using FISH and cytogenetic techniques. Examples of diseases that are diagnosed using FISH include Prader-Willi syndrome, Angelman syndrome, 22q13 deletion syndrome, chronic myelogenous leukemia, acute lymphoblastic leukemia, Cri-du-chat, Velocardiofacial syndrome, and Down syndrome. FISH on sperm cells is indicated for men with an abnormal somatic or meiotic karyotype as well as those with oligozoospermia, since approximately 50% of oligozoospermic men have an increased rate of sperm chromosome abnormalities. [19] The analysis of chromosomes 21, X, and Y is enough to identify oligozoospermic individuals at risk. [19]

    In medicine, FISH can be used to form a diagnosis, to evaluate prognosis, or to evaluate remission of a disease, such as cancer. Treatment can then be specifically tailored. A traditional exam involving metaphase chromosome analysis is often unable to identify features that distinguish one disease from another, due to subtle chromosomal features FISH can elucidate these differences. FISH can also be used to detect diseased cells more easily than standard Cytogenetic methods, which require dividing cells and requires labor and time-intensive manual preparation and analysis of the slides by a technologist. FISH, on the other hand, does not require living cells and can be quantified automatically, a computer counts the fluorescent dots present. However, a trained technologist is required to distinguish subtle differences in banding patterns on bent and twisted metaphase chromosomes. FISH can be incorporated into Lab-on-a-chip microfluidic device. This technology is still in a developmental stage but, like other lab on a chip methods, it may lead to more portable diagnostic techniques. [20] [21]

    Species identification Edit

    FISH has been extensively studied as a diagnostic technique for the identification of pathogens in the field of medical microbiology. [22] Although it has been proven to be a useful and applicable technique, it is still not widely applied in diagnostic laboratories. The short time to diagnosis (less than 2 hours) has been a major advantage compared with biochemical differentiation, but this advantage is challenged by MALDI-TOF-MS which allows the identification of a wider range of pathogens compared with biochemical differentiation techniques. Using FISH for diagnostic purposes has found its purpose when immediate species identification is needed, specifically for the investigation of blood cultures for which FISH is a cheap and easy technique for preliminary rapid diagnosis. [22]

    FISH can also be used to compare the genomes of two biological species, to deduce evolutionary relationships. A similar hybridization technique is called a zoo blot. Bacterial FISH probes are often primers for the 16s rRNA region.

    FISH is widely used in the field of microbial ecology, to identify microorganisms. Biofilms, for example, are composed of complex (often) multi-species bacterial organizations. Preparing DNA probes for one species and performing FISH with this probe allows one to visualize the distribution of this specific species within the biofilm. Preparing probes (in two different colors) for two species allows researchers to visualize/study co-localization of these two species in the biofilm and can be useful in determining the fine architecture of the biofilm.

    Comparative genomic hybridization Edit

    Comparative genomic hybridization can be described as a method that uses FISH in a parallel manner with the comparison of the hybridization strength to recall any major disruptions in the duplication process of the DNA sequences in the genome of the nucleus. [23]

    Virtual karyotype Edit

    Virtual karyotyping is another cost-effective, clinically available alternative to FISH panels using thousands to millions of probes on a single array to detect copy number changes, genome-wide, at unprecedented resolution. Currently, this type of analysis will only detect gains and losses of chromosomal material and will not detect balanced rearrangements, such as translocations and inversions which are hallmark aberrations seen in many types of leukemia and lymphoma.

    Spectral karyotype Edit

    Spectral karyotyping is an image of colored chromosomes. Spectral karyotyping involves FISH using multiple forms of many types of probes with the result to see each chromosome labeled through its metaphase stage. This type of karyotyping is used specifically when seeking out chromosome arrangements.


    Restricted License Requirements

    Practice as a clinical laboratory technologist within the areas of Cytogenetics Flow Cytometry/Cellular Immunology Histocompatibility Molecular Diagnosis to the extent such Molecular Diagnosis is included in Genetic Testing-Molecular and Molecular Oncology Molecular Diagnosis including but not limited to Genetic Testing-Molecular and Molecular Oncology for employment in cancer centers and designated training hospitals Stem Cell Process and Toxicology in New York State requires a license as a clinical laboratory technologist or a restricted license as a clinical laboratory technologist, unless otherwise exempt under the law.

    To receive a clinical laboratory technologist restricted license to practice in one of the previously mentioned areas in New York State you must:

    • be of good moral character
    • be at least 18 years of age
    • meet education requirements and
    • meet experience requirements.

    You must file an Application for a Restricted License (Form 1) and the other forms indicated, along with the appropriate fee, to the Office of the Professions at the address specified on each form. It is your responsibility to follow up with anyone you have asked to send us material.

    The specific requirements for licensure are contained in Title 8, Article 165 of New York's Education Law and Subpart 79-13 of the Regulations of the Commissioner of Education.

    You should also read the general licensing information applicable for all professions.

    The fee for a clinical laboratory technologist restricted license and first registration is $371.

    Fees are subject to change. The fee due is the one in law when your application is received (unless fees are increased retroactively). You will be billed for the difference if fees have been increased.

    • Do not send cash.
    • Make your personal check or money order payable to the New York State Education Department. NOTE: Your cancelled check is your receipt.
    • Mail your application and fee to:

    NYS Education Department
    Office of the Professions
    PO Box 22063
    Albany, NY 12201

    NOTE: Payment submitted from outside the United States should be made by check or draft on a United States bank and in United States currency payments submitted in any other form will not be accepted and will be returned.

    Partial Refunds

    Individuals who withdraw their licensure application may be entitled to a partial refund.

    • For the procedure to withdraw your application, contact the Clinical Laboratory Technology Unit by e-mailing [email protected] or by calling 518-474-3817 ext. 260 or by faxing 518-402-2323.
    • The State Education Department is not responsible for any fees paid to an outside testing or credentials verification agency.

    If you withdraw your application, obtain a refund, and then decide to seek New York State licensure at a later date, you will be considered a new applicant, and you will be required to pay the licensure and registration fees and meet the licensure requirements in place at the time you reapply.

    Education Requirements

    To meet the professional education requirements for a clinical laboratory technologist restricted license, you must present evidence of completion of a baccalaureate or higher degree program in the major of biology, chemistry, the physical sciences, or mathematics from a program registered by the State Education Department or determined by the Department to be the substantial equivalent.

    In addition to the degree requirement, you must complete an approved training program in the specific area in which you are seeking a restricted license. The content of the training program shall be described and attested to by the clinical director of the laboratory in which the program is located prior to the beginning of the your program* using the respective Form 4.

    *NOTE: You may not begin a program until the application has been approved and a certificate has been issued.

    The training program shall consist of not less than one year of full-time training in the specific area in which you are seeking certification, which shall consist of no less than 1750 hours in a calendar year, in the specific area in which you are seeking certification, or the part-time equivalent thereof, as determined by the department.

    The respective areas for each field are:

    Cytogenetics

    A program in cytogonetics must contain knowledge of:

    • chromosome structure/behavior and its correlation with phenotype and of chromosomal abnormalities.

    The program shall also include but need not be limited to:

    • general laboratory principles and skills
    • including infection control and aseptic technique quality control and quality assurance
    • clinical cytogenetics
    • general knowledge of human genetics
    • laboratory mathematics
    • the collection, handling, preparation and processing of pertinent specimens
    • the use of appropriate cell culture techniques
    • the principles and techniques for harvesting specimens or cell cultures and,
    • the principles and techniques of chromosome banding, staining, analysis, and instrumentation

    Flow Cytometry/Cellular Immunology

    A program in flow cytometry/cellular immunology must contain knowledge of:

    • The technique for counting, sorting, and characterization of cells suspended in a fluid stream based on their physical properties and expression of cell surface molecules

    The program shall also include but need not be limited to:

    • general laboratory principles and skills
    • infection control and aseptic technique
    • quality control and quality assurance
    • instrumentation and equipment
    • the basic principles of flow cytometry, including specimen preparation, fluidics and electronics
    • fluorochrome selection
    • antibody selection
    • the design of flow cytometry procedures, including routine standardization and quality management and
    • specific clinical applications.

    Histocompatibility

    A program in histocompatibility must contain knowledge of:

    • clinical immunology
    • immunogenetics
    • basic molecular biology and
    • laboratory mathematics.

    The program shall also include but need not be limited to:

    • general laboratory principles and skills, including infection control and aseptic technique
    • the practice of HLA typing and HLA antibody testing
    • specimen collection, processing and handling
    • instrumentation and equipment
    • reagent preparation and quality control
    • quality assurance, principles and techniques of histocompatibility assays, and crossmatching
    • antibody screening and identification and,
    • determination of degree of HLA matching.

    Molecular Diagnosis Restricted to Molecular Diagnosis Included In Genetic Testing-Molecular and Molecular Oncology

    A program in molecular diagnosis must contain knowledge of:

    • the role of molecular genetics in tumor diagnosis and individualized tumor therapies that are being defined and implemented.

    The program shall also include but need not be limited to:

    • general laboratory principles
    • infection control and aseptic technique
    • quality control and quality assurance
    • applicable laboratory skills
    • general principles of molecular biology, clinical molecular genetics and molecular diagnosis
    • laboratory mathematics
    • basic principles of nucleic acid extraction, modification, amplification, identification, and unidirectional workflow techniques to avoid cross contamination
    • electrophoresis and other separation techniques
    • transfer and hybridization techniques and specific techniques of nucleic acid amplification and identification.

    Molecular Diagnosis Not Restricted to Molecular Diagnosis Included in Genetic Testing-Molecular and Molecular Oncology for Employment in Cancer Centers and Designated Training Hospitals

    A program in molecular diagnosis must contain knowledge of:

    • the role of molecular genetics in tumor diagnosis and individualized tumor therapies that are being defined and implemented.

    The program shall also include but need not be limited to:

    • general laboratory principles
    • infection control and aseptic technique
    • quality control and quality assurance
    • applicable laboratory skills
    • general principles of molecular biology, clinical molecular genetics and molecular diagnosis
    • laboratory mathematics
    • basic principles of nucleic acid extraction, modification, amplification, identification, and unidirectional workflow techniques to avoid cross contamination
    • electrophoresis and other separation techniques
    • transfer and hybridization techniques and specific techniques of nucleic acid amplification and identification and
    • additional training in molecular diagnosis acceptable to the Department that would enable you to practice competently.

    Stem Cell Process

    A program in stem cell process must contain knowledge of:

    The program shall also include but need not be limited to:

    • general laboratory principles and skills
    • infection control and aseptic technique
    • instrumentation and equipment
    • quality control and quality assurance
    • laboratory mathematics
    • the process of handling stem cell specimens in the laboratory
    • enumeration and characterization of stem cells
    • ABO/Rh confirmatory typing and,
    • reagent preparation.

    Toxicology

    A program in toxicology must contain knowledge of:

    • laboratory methods in toxicology, including qualitative and quantitative determination of xenobiotics present in biological specimens.

    The program shall also include but need not be limited to:

    • general laboratory principles and skills
    • basic principles of chemistry, biology, and the physical sciences
    • basic principles of pharmacology
    • basic principles of purification, separation, and extraction techniques
    • instrumentation and equipment
    • quality control and quality assurance
    • laboratory mathematics
    • the principles of immunoassay techniques
    • preparation and processing of biological specimens for toxicological analysis
    • the principles of analytical techniques
    • review and certification of toxicology results and,
    • aseptic technique and infection control and specific clinical applications

    The Department must receive, directly from the clinical laboratory director of the program, verification of completion of an approved training program using the respective Form 4A.

    Additional Educational Requirements

    New York State Public Health Law and Regulations

    The laws, rules and regulations listed below can be accessed on the Web at www.wadsworth.org/regulatory/clep/laws

    • Article V, Title V Clinical Laboratory and Blood Banking Services
    • Article 31 Human Blood and Transfusion Services
    • Article 27F HIV and AIDS Related Information
    • Article 43-B, Organ, Tissue, and Body Parts
    • Article V, Title VI Laboratory Business Practices
    • Section 79.1 of the New York State Civil Rights Law, Confidentiality of Genetic Testing
    • Part 19 of 10 (NYCRR) Clinical Laboratory Directors
    • Subpart 34-2 of 10 (NYCRR) Laboratory Business Practices
    • Subpart 58-1 of 10 (NYCRR) Clinical Laboratories
    • Subpart 58-2 of 10 (NYCRR) Blood Banking
    • Subpart 58-5 of 10 (NYCRR) Hematopoietic Progenitor Cell Banks
    • Subpart 58-8 of 10 (NYCRR) Human Immunodeficiency Virus (HIV) Testing
    • Subpart 63 of 10 (NYCRR) AIDS/HIV Testing, Reporting and Confidentiality

    Federal Laws and Regulations

    The laws and regulations listed below can be accessed on the Web at www.cms.hhs.gov/clia/

    Requirements at a Glance

    You may print and keep this checklist as a reminder of what forms you need to file. This is for your reference and should not be submitted with your application forms. You should also keep a copy of all application forms submitted.


    Watch the video: Η ΓΕΝΕΤΙΚΗ ΚΑΤΑΓΩΓΗ ΤΩΝ ΕΛΛΗΝΩΝ (May 2022).