关键词: 实验兔, RAPD, 体系优化, 引物筛选

Abstract: Objective The optimal reaction system and primers of RAPD-PCR in experimental Rabbits were established. Methods One hundred and fifty samples were collected from WHBE rabbits, JW rabbits and NZW rabbits，and their DNA was extracted by standard procedure of phenol/chloroform. The optimization of its RAPD-PCR reaction system, the most appropriate concentration of Mg²⁺, dNTP, Taq polymerase, random primers and DNA template, was studied. Sixty primers were used to amplify DNA of experimental rabbits with RAPD-PCR methods. Results The optimal PCR system for RAPD analysis was carried out in 20 ul reaction volumes containing the following: 10×loading buffer 2ul, Mg2+ 1.5 mmol/L, dNTP 0.25 mmol/L, Taq polymerase 1.25 Units, random primer 5 jumol/L, DNA template 40 ng and sterile distilled water to total volume. On the basis of preliminary experiments, twenty-five primers with high polymorphism were selected in our study. Conclusion The selected 25 primers and optimalizing PCR reaction system can applied to RAPD reaction in experimental rabbit.

Key words: Experimental rabbit, RAPD, System optimization, Primer screening