POLIMERASE CHAIN REACTION (PCR)

POLIMERASE CHAIN REACTION (PCR)

• Kary Mullis (1984)• PENGGANDAAN FRAGMEN SPESIFIK DNA

SECARA INVITRO JUTAAN KOPI FRAGMEN DNA SECARA CEPAT DAN AKURAT

REPLIKASI

PCR

Basic Components

• TEMPLATE ATAU TARGET (FRAGMEN GEN YANG DIINGINKAN)

• DNA ATAU RNA

• dNTPs

• PRIMER

– OLIGONUKLETIDA PENDEK (15-30 BP)

– TERIKAT DENGAN SEKUENS SPESIFIK PADA TEMPLATE

• ENZIM

– HEAT-STABILE DNA POLIMERASE –Taq DNA POLIMERASE

– REVERSE TRANSCRIPTASE

Step 1 : Denaturation (separation of double-stranded DNA) by Heat (95° C)

Step 2 : Primer Anneals (50 - 65° C)

sstep 3 Primer Extension by Taq Polymerase (72° C)

End of the 1st PCR Cycle Results in 2 Copies of Target Sequence

CYCLE NUMBER AND AMPLICONS

VARIASI GENETIK (POLIMORFISME)

SINGLE NUCLEOTIDE POLYMORPHISMS (SNP) SPESIFIC REPEATED SEQUENCE (TANDEM REPEAT)