SNP - Single nucleotide polymorphism

1. SNPs and ESTs
Pruthvish R

2. • Single nucleotide polymorphisms (SNPs,
pronounced ‘snips’) are the most common
type of genetic variation amongst people.

3. 2007 Scientific Breakthrough of
the Year

4. SNPs
• A SNP is defined as a single base change in a DNA
sequence that occurs in a significant proportion
(more than 1 percent) of a large population.
• Each single nucleotide polymorphism represents
a difference in a single DNA base, A, C, G or T, in a
person’s DNA.
• On average they occur once in every 300 bases
and are often found in the DNA between genes.

5. • By definition, one map unit (m.u.) is equal to
one percent recombinant phenotypes.
• In honor of the work performed by Morgan,
one m.u. is also called one centimorgan (cM).

6. Facts about SNPs
►SNPs are found in
 coding and (mostly) noncoding regions.
►Occur with a very high frequency
 about 1 in 1000 bases to 1 in 100 to 300 bases.
►The abundance of SNPs and the ease with which they can be
measured make these genetic variations significant.
►SNPs close to particular gene acts as a marker for that gene.
►SNPs in coding regions may alter the protein structure made
by that coding region.

7. SNPs may / may not alter protein
structure

8. SNPs act as gene markers

9. SNP maps
►Sequence genomes of a large number of
people
►Compare the base sequences to discover
SNPs.
►Generate a single map of the human genome
containing all possible SNPs => SNP maps

11. SNP Profiles
• Genome of each individual contains distinct SNP
pattern.
• People can be grouped based on the SNP profile.
• SNPs Profiles important for identifying response
to Drug Therapy.
• Correlations might emerge between certain SNP
profiles and specific responses to treatment.

12. Techniques to detect known
Polymorphisms
• Hybridization Techniques
– Micro arrays
– Real time PCR
• Enzyme based Techniques
– Nucleotide extension
– Cleavage
– Ligation
– Reaction product detection and display

14. Techniques to detect unknown
Polymorphisms
• Direct Sequencing
• Microarray
• Cleavage / Ligation
• Electrophoretic mobility assays

15. SIGNIFICANCE OF SNPs
• IN DISEASE DIAGNOSIS
• IN FINDING PREDISPOSITION TO DISEASES
• IN DRUG DISCOVERY & DEVELOPMENT
• IN DRUG RESPONSES
• INVESTIGATION OF MIGRATION PATTERNS

16. Applications
• SNPs are used for identification and forensics
• SNPs are used for mapping and genome-wide
association studies of complex diseases
• SNPs are used for estimating predisposition to
disease
• SNPs are used for immigration & citizenship in the
UK
• SNPs are used to predict specific genetic traits
• SNPs are used for classifying patients in clinical
trials

17. Types of SNPs
• Noncoding SNPs
– 5’ UTR
– 3’ UTR
– Introns
– Intergenic Regions
– Pseudogenes
– Regulatory
• Splicing
• Transcriptional regulation (promoter & TF binding sites)
• Translational regulation (initiation or termination)
• Regulatory miRNA target sites
• Coding SNPs
– Synonymous SNPs (third position variation)
– Replacement SNPs (change Amino acid)
• Functional SNPs (acceptable amino acid replacement)
• Non-functional SNPs (traits & diseases)

18. EST
• Expression Sequence Tag
– Expressed sequence tags (ESTs) are short
sequence reads, typically within the range of 00–
700 bp, obtained from randomly selected cDNA
clones
• cost-effective approach for the rapid discovery
and characterization of expressed genes

19. • Purified mRNA is used as a template for reverse
transcriptase using either oligo(dT) as a primer
for the first-strand synthesis or, alternatively,
random hexamer primers.
• After nicking the RNA–DNA hybrid with RNAse H,
second-strand synthesis proceeds using the RNA
fragments as primers and DNA polymerase I for
the extension to construct a cDNA library.
• reverse transcriptase polymerase chain reaction
(RT-PCR)

20. • cDNA libraries prepared from various
organisms, tissues and cell lines using
directional cloning
• ESTs represent partial sequences of cDNA
clones (300 bp -> 700 bp)

21. Applications
• There are indispensable for gene structure prediction,
gene discovery and genome mapping:
-> provide experimental evidence for the position of
exons
-> provide regions coding for potentially new proteins
-> characterization of splice variants and alternative
polyadenilation
• Provide an alternative to library screening
-> short tag can lead to a cDNA clone
• Provide an alternative to full-length cDNA sequencing
-> sequences of multiple ESTs can reconstitute a full-
length cDNA
• Single Nucleotide Polymorphism (SNP) data mining

24. • The challenge associated with identifying
genes from genomic sequences varies
among organisms and is dependent upon
genome size as well as the presence or
absence of introns--the intervening DNA
sequences interrupting the protein coding
sequence of a gene