[目的]建立一套适合大竹蛏的AFLP分子标记反应体系,为大竹蛏的遗传多样性研究提供技术支持.[方法]采用比较实验法,以大竹蛏基因组DNA为模板,对其AFLP体系中的酶切时间、预扩产物稀释倍数及选扩引物E+3/M +3的配比等进行优化.[结果]优化的AFLP体系为:酶切时的DNA模板浓度为100 ng/μl,双酶切2h；预扩模板最佳用量1.0μl；预扩产物的最适稀释倍数为40倍;选扩引物浓度配比为1∶4.同时,筛选出12对引物组合,可用于大竹蛏的AFLP分析.[结论]采用优化的AFLP反应体系及筛选后的引物,对大竹蛏群体进行了初步扩增,得到了清晰的电泳图谱.%[Objective] The research aimed to establish the operation rules for AFLP-PCR reaction system in Solen grandis and provide basis for its application in genetic diversity. [ Method] With comparing experimentation, by using genomic DNA of Solen grandis adjusting the time for DNA double enzymes digestion reaction, the dilute multiple for the per-amplification production and different proportion in selective primers M +3/E +3 to select the optimization. [ Result] The optimum AFLP system was as followed: concentration of template DNA is 100 ng μl in the digest system and 2 hours for the reaction; 1.0 microlitre ligation products as the template of pre-amplification; 40 times of dilution for the products of pre-amplification with 1 ×TE for selective amplification; the ratio of primer concentration was 1:4. And 12 pairs of informative primers were identified and recommended for AFLP analysis of Solen grandis. [ Conclusion] The PCR amplification of Solen grandis were analyzed by using the optimized AFLP reaction system and most primers amplified clear bands.
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