从茶树EST序列中搜寻出候选SNPs位点,在候选SNPs两侧设计测序引物进行PCR扩增,并对扩增产物双向测序验证;然后从每个扩增子中选择一个确证的SNPs,设计dCAPS引物,并进行扩增和酶切验证,将SNPs转化为dCAPS标记;最后,在长叶白毫×福鼎大白的F1群体中检测了标记的分离情况。结果发现,20对测序引物中有11对扩增成功,共确证了17个SNPs,占检测总数的54.8%;在设计的11对dCAPS引物中,有8对在长叶白毫、福鼎大白和龙井43等8个品种中表现出多态性,成功转化为dCAPS标记,转化成功率72.7%;8个dCAPS标记中的DCC229和DCC371在长叶白毫和福鼎大白之间表现有多态性,经检测,它们在这2个品种的F1群体中均符合孟德尔1︰1分离。
SNPs were mined from tea-derived EST sequences. Primers were designed to evaluate the predicted SNPs in genomic DNA from tea plant. The PCR products were sequenced from both directions to confirm the valid SNPs. Meanwhile, the dCAPS primers were designed based on the valid SNPs and the genomic DNA was used for amplification. The amplicons were digested by restrict enzymes to convert SNPs into dCAPS markers. Furthermore, we also detected the separation of the new markers in a F1 population. The results showed that seventeen SNPs (54.8%) were confirmed in 11 amplycons, eight SNPs were successfully converted to dCAPS markers (72.7%), two of them which have polymorphisms in parents exhibited a separate ratio of 1︰1 in the F1 population. The new markers would be useful in tea genetic research. It is also believed that dCAPS technology will promote the use of SNP technology in genetics and breeding research in tea plant.
Key words：
tea; EST; SNP; dCAPS
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