Sts

STS stands for sequence tagged site which is short DNA sequence, generally between 100 and 500 bp in length, that is easily recognizable and occurs only once in the chromosome or genome being studied. 

1. Sequence Tagged Sites Nosheen A fzal BI141014

2. Sequence Tagged Sites A  sequence tagged site or STS is simply a short  DNA sequence , generally between 100 and 500  bp  in length, that is easily recognizable and occurs only once in the chromosome or genome being studied.  

3. History The STS concept was introduced by Olson et al (1989). In assessing the likely impact of the Polymerase Chain Reaction (PCR) on human genome research, they recognized that single-copy DNA sequences of known map location could serve as markers for genetic and physical mapping of genes along the chromosome. 

4. Why? STSs can be easily detected by the polymerase chain reaction (PCR) using specific primers. For this reason they are useful for constructing genetic and physical maps from sequence data reported from many different laboratories. serve as landmarks on the developing  physical map of a genome.

5. Process of Mapping STS To map a set of STSs a collection of overlapping DNA fragments from a single chromosome or the entire genome is required. To do this, the genome is first broken up into fragments. The fragments are then replicated up to 10 times in bacterial cells to create a library of DNA clones. The polymerase chain reaction (PCR) ?  is then used to determine which fragments contain STSs. Special primers ?  are designed to bind either side of the STS to ensure that only that part of the DNA is copied. 

6. Mapping Process of STS

7. Types of STS Marker STS include such markers as microsatellites (SSRs, STMS or SSRPs ), SCARs, CAPs , and ISSRs .

8. Microsatellites Polymorphic loci present in nuclear DNA and organellar DNA that consist of repeating units of 1-10 base pairs, most typically, 2-3 bp in length, also called Simple Sequence Repeats (SSR), Sequence-Tagged Microsatellite Sites (STMS) or Simple Sequence Repeats Polymorphisms (SSRP). SSRs are highly variable and evenly distributed throughout the genome. This type of repeated DNA is common in eukaryotes.

9. Working These polymorphisms are identified by constructing PCR primers for the DNA flanking the microsatellite region. The flanking regions tend to be conserved within the species, although sometimes they may also be conserved in higher taxonomic levels . They are widely used for DNA profiling in kinship analysis (especially paternity testing) and in forensic identification. They are also used in genetic linkage  analysis/marker assisted selection to locate a gene or a mutation responsible for a given trait or disease. Microsatellites are also used in population genetics to measure levels of relatedness between subspecies, groups and individuals .

10. Sequence Characterized Amplified Region (SCAR) DNA fragments amplified by the Polymerase Chain Reaction (PCR) using specific 15-30 bp primers, designed from nucleotide sequences established in cloned RAPD (Random Amplified Polymorphic DNA) fragments linked to a trait of interest. By using longer PCR primers, SCARs do not face the problem of low reproducibility generally encountered with RAPDs . Obtaining a co-dominant marker may be an additional advantage of converting RAPDs into SCARs.

11. Strengths The main advantage of SCARs is that they are quick and easy to use. In addition, SCARs have a high reproducibility and are locus-specific. Due to the use of PCR, only low quantities of template DNA are required. Weaknesses Disadvantages include the need for sequence data to design the PCR primers. Applications SCARs are locus specific and have been applied in gene mapping studies and marker assisted selection.

12. Cleaved Amplified Polymorphic Sequences (CAPS) Cleaved Amplified Polymorphic Sequences (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.

13. Developing CAPS markers Sequence the RFLP probe. Design primers to amplify 800–2,000-bp DNA fragments. Targeting introns or 3' untranslated regions should increase the chance of finding polymorphisms The PCR product is cloned and sequenced. PCR amplify DNA fragments from target genotypes, separately digest the amplicons with one or more restriction enzymes. Screen the digested amplicons for polymorphism on gels stained with ethidium bromide.

14. Advantages of CAPS co-dominant and locus-specific. CAPS genotypes are easily scored and interpreted. Markers are easily shared between laboratories. It does not require the use of radioactive isotopes, and it is more amenable, therefore, to analyses in clinical settings.

15. Inter-simple Sequence Repeats (ISSRs )  ISSRs are amplified by PCR using microsatellite core sequences as primers with a few selective nucleotides as anchors into the non-repeat adjacent regions (16-18 bp ). About 10-60 fragments from multiple loci are generated simultaneously, separated by gel electrophoresis and scored as the presence or absence of fragments of particular size. 

16. Strengths The main advantage of ISSRs is that no sequence data for primer construction are needed. Because the analytical procedures include PCR, only low quantities of template DNA are required. Furthermore, ISSRs are randomly distributed throughout the genome . Weaknesses Because ISSR is a multilocus technique, disadvantages include the possible non-homology of similar sized fragments. Moreover, ISSRs, like RAPDs, can have reproducibility problems .

17. Applications Because of the multilocus fingerprinting profiles obtained, ISSR analysis can be applied in studies involving genetic identity, parentage, clone and strain identification, and taxonomic studies of closely related species. In addition, ISSRs are considered useful in gene mapping studies.

18. Applications of STS STSs are very helpful for: detecting microdeletions in some genes. For example, some STSs can be used in screening by PCR to detect microdeletions in Azoospermia (AZF) genes in infertile men . Identification of genes in elephants could provide additional information for evolutionary studies and for evaluating genetic diversity in existing elephant populations.

19. Reference Marwal , Avinash , Anurag Kumar Sahu , and R.K. Gaur. "Molecular Markers."  Animal Biotechnology  (2014): 289-305. Web. 25 Oct. 2017. https://www.ncbi.nlm.nih.gov/probe/docs/techsts/