BIOL 142 Chapter Notes - Chapter 20.2: Polymerase Chain Reaction, Tandem Repeat, Dna Profiling
30 Apr 2018
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20.2$The$Polymerase$Chain$Reaction$
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Polymerase*chain*reaction*(PCR):!a!laboratory!technique!for!rapidly!generating!millions!of!identical!
copies!of!a!specific!stretch!of!DNA;!works!by!multiple!rounds!of!DNA!replication,!starting!from!primers!
designed!by!an!investigator!to!amplify!particular!DNA!sequences!
• Another!way!to!make!identical!copies!of!a!region!of!DNA!
• An!in!vitro!DNA!synthesis!reaction!that!uses!a!DNA!polymerase!to!replicate!a!specific!section!of!
DNA!
!!
Requirements!of!PCR!
• Only!possible!when!a!researcher!already!has!some!information!about!DNA!sequences!that!
surround!the!DNA!in!question!
o Needed!because!you!have!to!start!by!synthesizing!short!lengths!of!single-stranded!DNA!that!
match!sequences!on!either!side!of!the!region!of!interest!and!act!as!primers!
• Works!by!repeating!three!steps!over!and!over!
o Each!repeat!is!called!a!cycle!
o Each!cycle!doubles!the!number!of!target!sequences!
o Doubles!because!each!strand!of!DNA!synthesized!in!a!previous!cycle!serves!as!a!
template!in!the!next!cycle!
• For!the!wide!array!of!organisms!with!completely!sequenced!genomes,!researchers!can!easily!find!
primer!sequences!to!use!in!cloning!almost!any!target!gene!by!PCR!
!!
DNA*fingerprinting*(DNA*profiling/DNA*typing):!!any!of!several!methods!for!identifying!individuals!by!
unique!features!of!their!genomes;!commonly!involves!using!PCR!to!produce!many!copies!of!short!
tandem!repeats!(microsatellites)!and!then!analyzing!their!lengths!
• Tandem*repeat*sequences:!short!DNA!sequences!that!are!repeated!one!after!another!along!part!
of!a!chromosome;!!vary!in!repeat!number!between!individuals;!the!core!of!modern!DNA!
fingerprinting!
o Many!eukaryotic!genomes!have!thousands!of!loci!sites!that!contain!tandem!repeats!
• 2!categories!of!tandem!repeats!
o Short*tandem*repeats*(STRs)/microsatellites/simple*sequence*repeats*(SSRs):!short!(2-6!
base!pair)!DNA!sequence!that!is!repeated!sequentially!along!a!chromosome!
o Most!often!used!in!fingerprinting!because!of!their!shorter!size!
o Shorter*size*makes*them*easier*to*amplify*by*PCR!
o Thought!to!originate!when!DNA!polymerase!skips!or!mistakenly!adds!extra!bases!
during!replication!
o STRs!are!hypervariable:!vary!among!individuals!far!more!than!other!types!of!
sequences!
o Each!repeat!number!variant!of!an!STR!(aka!STR!allele)!is!transmitted!from!parent!to!
offspring!like!any!other!DNA!sequence!
o Minisatellites/variable*number*tandem*repeats*(VNTRs):!a!noncoding!stretch!of!eukaryotic!
DNA!consisting!of!a!repeating!sequence!that!is!from!8!to!about!10!base!pairs!long!
• PCR!allows!minute!amounts!of!DNA!to!be!analyzed!!
o STR!is!amplified!
o Gel!electrophoresis!is!used!to!determine!the!number!of!repeats!
o Primers!allow!the!analysis!of!many!different!STRs!in!a!single!polymerase!chain!reaction!
• Helpful!in!parental!analysis!and!criminal!investigation!
