Analisis daerah d-loop dna mitokondria pada kerbau belang (bubalus bubalis) dengan menggunakan enzim restriksi (pcr- rflp)

Abstract

Spotted buffalo is slaughtered buffalo for sacrifices in Tana Toraja funeral ceremonies. Spotted buffalo is believed gives security in the hereafter. Therefore at the funeral ceremony, many spotted buffaloes slaughtered. Generally, hundreds of buffaloes from every class sacrificed every year. Further studies about DNA genetic pattern from spotted buffalo is needed in order to kept it existence. One of them is mitochondrial DNA expecially D-loop region. The objective of this research was to know the the connection between D-loop fragment variation and skin color variation in B. bubalis as a basic information to kept the existence of spotted buffalo. Tha research was performed using PCR-RFLP technique. DNA amplification in Dloop region used spesific primers CTDF and CTDR was then cutted using restriction enzymes Hae III, Alu I, Rsa I, Hinc II, Bam HI and Eco RI. PCR amplification of D-loop generated same size DNA bands that were approximately 1361 bp. Cutting PCR product with Hae III, Alu I, Rsa I, and Hinc II generated DNA bands with same pattern (monomorphic). Meanwhile Bam HI and Eco RI did not generated any cutted DNA band. Cutting with Hae III generated DNA bands those were 636 bp, 166 bp, and 559 bp. Cutting with Alu I generated DNA bands those were 71 bp, 82 bp, 365 bp, 191 bp, 276 bp, 185 bp, dan 191 bp. Cutting with Rsa I generated DNA bands those were 284 bp dan 1077 bp. Cutting with Hinc II generated DNA bands those were 1203 bp dan 158 bp. Restriction site in D-loop fragmen in buffalo had no polymorphism. The prediction is first it was caused by the lack of restriction enzymes that have been used. Second it can be caused by restriction enzymes cutting site that being used were at the non hypervariable D-loop region. Third it can be caused by buffalo domestication which centered in Indonesia or because of buffalo introduction to Indonesia from one distribution source. So it can be said that those buffaloes came from the same ancestor. Because the result was monomorphic, it can be concluded that there was no clear connection between mitochondrial variation in D-loop fragmen with spotted skin color in buffalo.