Tuesday, 27 April 2021

STS(Sequence Tagged Site) Mapping - Principle, Steps and Applications

  STS (Sequence Tagged Site) is a short sequence (200 to 500 base pair). That is unique in the genome. Its location and the sequence is known and therefore, it can be used as a landmark in genome mapping.

STS Mapping

   These STSs can be detected using amplification by PCR (Polymerase Chain Reaction) and are useful for localising mapping and sequence data.
  STS concept was given by olsen in 1999, in analysing the impact of PCR on human genome research. They found that single copy of DNA sequence of non map location could serve as a marker for genetic and physical mapping of gens along the chromosome.

EST (expressed sequence tag) :

  Sequence Tag Site derived from a region of DNA that is expressed in transcribed into mRNA is called EST.
• Purified mRNA is used to generate the corresponding cDNA (Complementary DNA) by reverse transcriptase.
• EST is the commonly used STS marker which is unique and produced by partial Sequencing of cDNA library clones.

  Since STS sequence may contain repetitive elements, sequence that appear elsewhere in the genome they posses market sequences at both ends of the site(STS) that are unique and conserved.
• STS include markers as microsatellites -
SSR : Simple Sequence Repeats
STMs : Sequence Tag Microsatellite Sites
SSRPs : Simple Sequence Repeat Polymorphism
CAPs : Cleaved Amplified Polymorphic Sequence
ISSRs : Inter Simple Sequence Repeats.

STS Mapping :

  A map representing the order and spacing of sequence tagged site, which in strength of DNA is known as STS mapping.
  STS mapping is essentially the same as map. Distances determined by linkage analysis except that the map distance is based on the frequency at which occurs between two markers.

Principle of STS Mapping :

To Map STS or ESt sites, a collection of chromosome fragments is examined for the presence of each sts for EST. 
Fragments may be derived from a single chromosome as the whole genome.
   Neighbouring STSs will tend to be on the same fragment where as those very far apart will only rarely be found on the same fragment.

Steps in STS Content Mapping

1) Fragmentation of genome -   

  Fermentation of the genome into overlapping plants is usually done by using rare cutting enzymes (restriction enzyme) to produce large DNA fragments.

2). Preparation of large fragment library -

  The fragments does generated are ligated to clone vectors of higher capacity like YAC.

3). Overlapping clone identification -

Two strategies are used to identify overlapping clones.
a) Clone Fingerprinting - Overlapping clones are identified based on common patterns of Restriction Enzyme fragments or repeat conserved between clones.
b) Chromosome walking -  
Overlapping clones are identified based on hybridization of two overlapping to a particular STS marker.

4) Preparation of SDS Content Map -

Arranging the overlapping fragments based on the presence or absence of a single particular probe.

Applications of STS mapping

•  STS content mapping is one of the best physical mapping methods by which high resolution is achieved.
•  STS based PCR produces a simple and reproducible pattern on agarose gel electrophoresis or polyacrylamide gel electrophoresis. In most case STS markers are Co-dominant i.e. allow heterozygotes to be distinguished from homeozygotes.


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