Limitations of RFLP

•   Requires high quantity and quality of DNA

•   Low polymorphism

•   Requires development of specific probe libraries

•   Requires radioactively labeled probes

 

Laborious and time consuming

6-5.3(a).1.2 Random amplification of polymorphic DNA (RAPD)

►It is the first molecular marker for PCR based genetic-mapping as well as DNA-fingerprinting.

In the field of genetic mapping PCR-based markers are considered as second generation molecular markers.

►RAPD markers are short of length (approx10nucleotides only)

►RAPD needs only one primer instead of a set of primers for amplification.

►It does not require any prior information about the DNA sequence of the desired organism.

►Polymorphism i.e. relatively variable DNA sequences between different species,occurs due to mutation or rearrangements either at or in between the primer binding sites.

►PCR products are represented by electrophoresis and visualized by ethidium bromide staining.

►It is a quick, simple and efficient technique as it does not involve blotting or hybridization steps.

►Requires only small amount of DNA (10 ng/reaction) and the process can be automated.

►Primers are non species specific and can be universal.

►RAPD products can be cloned, sequenced and converted to other types of markers (SCAR,SNP)