Molecular Bio Review

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Unformatted text preview: to PCR analysis FTATM paper Adv. = stable for many years/reusable/no quantification Disadv. = leapfrogging punches Differential Adv. = separates female epithelial cells from male sperm Disadv. = does not work with azoospermic semen Too much DNA complicates analysis Too little generates an incomplete profile Degraded DNA prevents amplification from larger loci / Proper collection and preservation of DNA is critical more on this later! Summary of the Process of DNA Typing with STRs %LRORJ\ 6DPSOH 2EWDLQHG IURP &ULPH 6FHQH RU 3DWHUQLW\ ,QYHVWLJDWLRQ '1$ '1$ ([WUDFWLRQ ([WUDFWLRQ '1$ '1$ 4XDQWLWDWLRQ 4XDQWLWDWLRQ 7HFKQRORJ\ 6HSDUDWLRQ DQG 'HWHFWLRQ RI 3&5 3URGXFWV 675 $OOHOHV 6DPSOH *HQRW\SH 'HWHUPLQDWLRQ 3&5 $PSOLILFDWLRQ 3&5 $PSOLILFDWLRQ RI 0XOWLSOH 675 PDUNHUV RI 0XOWLSOH 675 PDUNHUV Polymerase Chain Reaction In vitro preferential amplification of a specific segment of DNA from template. Amplify specific segment of DNA via primers. Taq DNA polymerase, MgCl2, Buffer, dNTPs, DNA template. *HQHWLFV &RPSDULVRQ RI 6DPSOH *HQRW\SH WR 2WKHU 6DPSOH 5HVXOWV *HQHUDWLRQ RI &DVH 5HSRUW ZLWK 3UREDELOLW\ RI 5DQGRP 0DWFK ,I PDWFK RFFXUV FRPSDULVRQ RI '1$ SURILOH WR SRSXODWLRQ GDWDEDVHV Butler, Figure 1.2 %XWOHU )LJXUH DNA in the Cell Cycling Conditions chromosome cell nucleus Denaturation, Annealing, Extension Double stranded DNA molecule Target Region for PCR Individual nucleotides %XWOHU )LJXUH PCR Copies DNA Exponentially through Multiple Thermal Cycles Original DNA target region Thermal cycle In 32 cycles at 100% efficiency, 1.07 billion copies of In 32 cycles at 100% efficiency, 1.07 billion copies of targeted DNA region are created targeted DNA region are created %XWOHU )LJXUH http://www.cstl.nist.gov/biotech/strbase/ Resulting Products Types of STR Repeats Microsatellite = simple sequence repeat (SSR) = short tandem repeat (STR) PCR can generate copies of a desired gene, or... Copies of a non-coding segment of DNA Short Tandem Repeats = STR Dinucleotide Trinucleotide Tetranucle...
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This note was uploaded on 09/21/2009 for the course MCB 3416 taught by Professor Ladd during the Fall '08 term at UConn.

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