林旻, 杨绍华, 林艳, 王月, 胡太蛟, 宋亚娜, 潘雷博, 陈在杰. 一种适用于高通量基因分型的水稻种子切片DNA制备技术[J]. 福建农业科技, 2023, 54(10): 40-47. DOI: 10.13651/j.cnki.fjnykj.2023.10.007
    引用本文: 林旻, 杨绍华, 林艳, 王月, 胡太蛟, 宋亚娜, 潘雷博, 陈在杰. 一种适用于高通量基因分型的水稻种子切片DNA制备技术[J]. 福建农业科技, 2023, 54(10): 40-47. DOI: 10.13651/j.cnki.fjnykj.2023.10.007
    shu
    LIN Min, YANG Shao-hua, LIN Yan, WANG Yue, HU Tai-jiao, SONG Ya-na, PAN Lei-bo, CHEN Zai-jie. Preparation Technique of DNA Extracted from the Slices of Rice Seeds Suitable for the High-throughput Genotyping[J]. Fujian Agricultural Science and Technology, 2023, 54(10): 40-47. DOI: 10.13651/j.cnki.fjnykj.2023.10.007
    Citation: LIN Min, YANG Shao-hua, LIN Yan, WANG Yue, HU Tai-jiao, SONG Ya-na, PAN Lei-bo, CHEN Zai-jie. Preparation Technique of DNA Extracted from the Slices of Rice Seeds Suitable for the High-throughput Genotyping[J]. Fujian Agricultural Science and Technology, 2023, 54(10): 40-47. DOI: 10.13651/j.cnki.fjnykj.2023.10.007
    shu

    一种适用于高通量基因分型的水稻种子切片DNA制备技术

    Preparation Technique of DNA Extracted from the Slices of Rice Seeds Suitable for the High-throughput Genotyping

      摘要
    • 摘要: 建立一种高质量、低成本的水稻种子切片DNA制备方法,用于育种群体的高通量基因分型。通过对比TPS法提取水稻叶片DNA及96孔板结合磁珠法提取切片种子DNA的纯度和浓度,进一步采用STARP(Semi-thermal asymmetric PCR)标记检测验证96孔板结合磁珠法提取水稻切片种子DNA的质量。结果表明:96孔板结合磁珠法提取水稻种子切片的DNA质量和纯度优于TPS法提取水稻叶片的DNA质量;STARP标记检测验证结果显示96孔板结合磁珠法提取水稻切片种子DNA可适用于GS3、Wx的STARP标记基因分型,其鉴定结果与测序结果一致。研究显示96孔板结合磁珠法可以从微量水稻种子切片中提出高回收率、高纯度、高完整度、无PCR抑制物的DNA用于高通量STATP法基因单倍型分析,可获得一种96孔板结合磁珠法提取DNA的高通量基因分型操作流程。

       

      Abstract: The preparation method of DNA with high quality and low cost extracted from the slices of rice seeds was established for the high-throughput genotyping of breeding populations. The degree of purity and concentration of DNA extracted from the leaves of rice by TPS method and those of DNA extracted from the slices of rice seeds by using the method of 96-well plates combined with magnetic beads were compared in this paper. Then, the quality of DNA extracted from the slices of rice seeds by using the method of 96-well plates combined with magnetic beads was further verified by STARP (Semi-thermal asymmetric PCR) marker detection. The results showed that the quality and the degree of purity of DNA extracted from the slices of rice seeds by using the method of 96-well plates combined with magnetic beads were better than those of DNA extracted from the leaves of rice by TPS method. The verification results of STARP marker detection showed that the method of 96-well plates combined with magnetic beads to extract the DNA from the slices of rice seeds could be applied to the STARP marker genotyping of GS3 and Wx, and the identification results were consistent with the sequencing results. The results showed that the method of 96-well plates combined with magnetic beads could be used to extract the DNA with high recovery rate, high purity, high integrity and no PCR inhibitors from the trace slices of rice seeds for the analysis of gene haplotype with the high-throughput STATP method. Therefore, the operation flow of high-throughput genotyping for the DNA extraction by using the method of 96-well plates combined with magnetic beads could be obtained.

       

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