PCR- Restrictionfragmentlengthpolymorphism(RFLP)

Students’presentation1By

FatimaZahran &Hanan Mahmoud

Principleofthetest

• PCR-RFLP is a techniquethat combines the PCRamplification with thedigestion of PCR productsby restriction enzymes(RE).

• Cleaving DNA intofragments of certainsizes, whose analysis onagarose gel results indifferent patterns offragment sizes, enablingthe identification.

First:Leishmania PCRanalysis

DNAextractionfrombloodsamples

ITS1PCRassay

Forspeciesidentification

PCRproductpurification(Qiagene kit)

RFLPusing

Hae IIIenzyme

M LT LM M LT LM

Buttherearesomedisadvantagesforthistechnique

• It does not detect all sequence and lengthvariations within or among amplicons, sincethe restriction enzymes used only recognise asmall number of potentially variable sites.

• Additionally, it requires large amount ofhighly pure DNA