Chapter 8 STRs

      What is needed in order to analyze STRs

o   Identify the flanking regions of the STR

o   Design primers that will bind to flanks

o   Which STR repeats are used most often & why

  Tetranucleotide repeats are used the most

  4 bp spread allows for better resolution in heterozygotes

What is stutter

  Stutter is a known biological artifact that results from PCR

template strand slippage causes stutter

      Amplicons that are 1 repeat unit less than the true allele

Simple repeats

contain units of identical length and sequence

(GATA)(GATA)(GATA)

Simple repeats with non-consensus alleles

contain an incomplete repeat unit (e.g., TH01 9.3)

Compound repeats

comprise two or more adjacent simple repeats

(GATA)(GATA)(GACA)

Complex repeats

contain several repeat blocks of variable unit length

      Allelic ladders   How they are made

  Created by combining locus-specific STR products from multiple individuals in a population

  Samples are co-amplified to produce an artificial sample containing the common STR alleles

what is the purpose of allelic ladders

Run with the test samples and provide a reference DNA size for each allele in the ladder

      Advantages for STR markers

o   Small product sizes are generally compatible with degraded DNA and PCR enables recovery of information from small amounts of material

o   Multiplex amplification with fluorescence detection enables high power of discrimination in a single test

o   Commercially available in an easy to use kit format

o   Uniform set of core STR loci provide capability for national and international sharing of criminal DNA profiles

o   Five-dye detection

  4 used to label PCR products

  1 used for the internal size standard

Mobility-modifying nucleotide linkers

  Made of hexaethyleneoxide (HEO) that shifts products 2.5 nt for each HEO unit

  Added to the 5’ end of the PCR primer so the product contains the extra molecules at one end

  Allows for the better separation of products whose sizes normally overlap

  Easier than changing the PCR primer binding site (done by Promega)

      Changes to CODIS

  CODIS core loci increased to 20

  Reduce the potential of random matches occurring within the dataset

  More and more samples are constantly being added

  Increases discrimination power in missing person investigations

  Encourages international data sharing efforts by having more loci in common with other countries for comparison purposes

Dys391

helps to confirm male

located on long arm of y-chromosome

only 2 null alleles found

Y-indel

helps with degraded samples

insertion (2) or deletion (1)

      Sex identification with Amelogenin

      AMELX on X chromosome

      AMELY on Y chromosome are homologous

  AMEL null mutations

      Only AMELX  or AMELY is detected in males

      No result in females

      Caused by

o   Deletions in the short arm

o   Primer binding site mutations

      STR base: public info on STRs

  References related to DNA testing and STRs

  Microvariant (off-ladder) alleles

  Triallelic patterns

  PowerPoints & articles to aid in understanding STRs

  STR fact sheets

  Review of past & current technology

  Primer sequences

  Population data for STRs