Rapid and sensitive identification of buffalo's, cattle's and sheep's milk using species-specific PCR and PCR-RFLP techniques

Abstract

For the rapid, specific and sensitive identification of buffalo's, cattle's and sheep's milk, species-specific PCR and PCR-RFLP techniques were developed. DNA from small amount of fresh milk (100 μL) was extracted to amplify the gene encoding species-specific repeat (SSR) region and the mitochondrial DNA segment (cytochrome-b gene). PCR amplification size of the gene encoding SSR region was 603 bp in both buffalo's and cattle's milk, while in sheep's milk was 374 bp. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique was used to discriminate between buffalo's and cattle's milk. Restriction analysis of PCR-RFLP of the mitochondrial cytochrome-b segment (359 bp) analysis showed difference between buffalo's and cattle's milk. Where, the fragment length (bp) generated by TaqI PCR-RFLP were 191 and 168, whereas no fragments were obtained in cattle's milk for cytochrome-b gene (359 bp). The proposed PCR and PCR-RFLP assays rep resent a rapid and sensitive method applicable to the detection and authentication of milk species-specific. © 2006 Elsevier Ltd. All rights reserved.