谷子种子DNA快速提取新方法
|
摘要
为满足谷子高通量分子标记检测的需要,探究在不使用液氮研磨情况下,无需经过发芽,改进CTAB方法,直接从干种子中提取DNA。结果显示,采用新方法对谷子种子提取的DNA经过分光光度计以及琼脂糖电泳检测得出,A_(260/280)值在1.88~1.92,质量浓度在1 662.85~2 551.89 ng/μL;并以此为模板进行SSR-PCR扩增,结果显示,所得条带清晰、多态性丰富。该方法提取的谷子种子DNA质量稳定,能够满足分子标记检测的要求,同时简化了操作流程,实现了谷子种子DNA的高通量快速提取,为谷子高通量分子标记检测奠定了基础。
To meet the need of high-throughput molecular marker detection of millet, millet seeds were used as materials to explore DNA extraction of millet seeds directly by simplified CTAB without using liquid nitrogen grinding and without germination. The results showed that the DNA extracted from the seeds of millet using the new method was detected by spectrophotometer and agarose gel electrophoresis, the A_(260/280) value was 1.88-1.92, and the mass concentration was 1 662.85-2 551.89 ng/μL. Using this as a template SSR-PCR amplification, the resulting bands was clear, polymorphisms was rich. The DNA quality of millet seeds extracted by the method is stable and can meet the requirements of molecular marker detection. At the same time, the operation process is simplified, and the high throughput and rapid extraction of DNA in millet seed is realized, which lays a foundation for high flux molecular marker detection of millet.
To meet the need of high-throughput molecular marker detection of millet, millet seeds were used as materials to explore DNA extraction of millet seeds directly by simplified CTAB without using liquid nitrogen grinding and without germination. The results showed that the DNA extracted from the seeds of millet using the new method was detected by spectrophotometer and agarose gel electrophoresis, the A_(260/280) value was 1.88-1.92, and the mass concentration was 1 662.85-2 551.89 ng/μL. Using this as a template SSR-PCR amplification, the resulting bands was clear, polymorphisms was rich. The DNA quality of millet seeds extracted by the method is stable and can meet the requirements of molecular marker detection. At the same time, the operation process is simplified, and the high throughput and rapid extraction of DNA in millet seed is realized, which lays a foundation for high flux molecular marker detection of millet.
引文
[1]周晶,曾庆涛,刘铨义,等.一种适用于棉花种子的DNA快速提取方法[J].作物杂志,2014(2):31-33.
[2]魏琦超,畅丽萍,周岩,等.利用改良CTAB法提取小麦干种子总DNA[J].山西农业科学,2009,37(6):30-32.
[3]王惠,郭峰,关超,等.适用于SSR分析的半粒水稻干种子DNA快速提取[J].科技导报,2013(25):58-60.
[4]田丽波,谷幸幸,商桑,等.苦瓜基因组DNA的提取及ISSR扩增体系的优化[J].中国农学通报,2013,29(4):88-93.
[5]韩玉杰,贾炜珑,王自霞,等.几种提取植物DNA方法的比较[J].山西农业科学,2008,36(7):17-19.
[6]凌莉,李志勇,黄韵,等.食品中植物基因组DNA提取纯化方法研究进展[J].食品科技,2012(5):6-10,15.
[7]张换样,李静,南芝润,等.甘薯DNA的小量快速提取[J].山西农业科学,2009,37(1):12-14.
[8]戴剑,洪德林,张大栋,等.一种快速高效的DNA提取方法研究[J].麦类作物学报,2011(3):437-442.
[9]龚双军,杨立军,刘辉,等.1种小麦白粉病菌DNA基因组的微量简捷提取方法[J].微生物学杂志,2011(1):24-27.
[10]颜松.药用植物基因组DNA提取及铁皮石斛RAPD反应体系的优化[D].重庆:西南交通大学,2009.
[11]张晓祥,王玲,寿路路.一种快速提取小麦基因组DNA的改良CTAB方法[J].中国农学通报,2012,28(36):46-49.
[12]许明,程祖锌,黄志伟,等.一种适于转基因水稻PCR检测的微量DNA快速提取法[J].生物技术通报,2010(3):128-130.
[13]王兰,龙云铭,刘耀光.一种用于PCR的植物基因组DNA快速制备方法[J].分子植物育种,2009,7(2):425-428.
[14]楼巧君,陈亮,罗利军.三种水稻基因组DNA快速提取方法的比较[J].分子植物育种,2005(5):749-752.
[15]任海龙.药用植物大黄种子基因组DNA的提取方法研究[J].山西农业科学,2018,46(3):325-327,349.
[16]刘风路,张金丽,李靖,等.西南地区榧树属植物3种不同总DNA提取方法的比较分析[J].天津农业科学,2018,24(3):1-4.
[17]文静,谢培,孙琛,等.珠子参新鲜和干燥块茎DNA提取方法的比较研究[J].陕西农业科学,2018,64(2):1-3,11.
[18]盛鸥,张玉娥,邓贵明,等.香蕉不同组织的基因组DNA制备方法的比较[J].分子植物育种,2017,15(12):5052-5059.
[19]董永军,王陆军,郝建平,等.玉米干种子基因组DNA提取方法的改进[J].山西农业科学,2017,45(12):1903-1906.
[20]王瑞云,刘笑瑜,王海岗,等.用高基元微卫星标记分析中国糜子遗传多样性[J].中国农业科学,2017,50(20):3848-3870.
[21]闫玖英,马长青,常博,等.改良CTAB法用于苹果果实基因组DNA的提取[J].分子植物育种,2017,15(9):3610-3615.
[22]曾巧英,凌秋平,胡斐,等.5种提取方法对甘蔗汁中DNA提取的效果[J].安徽农业科学,2017,45(25):136-138,152.
[2]魏琦超,畅丽萍,周岩,等.利用改良CTAB法提取小麦干种子总DNA[J].山西农业科学,2009,37(6):30-32.
[3]王惠,郭峰,关超,等.适用于SSR分析的半粒水稻干种子DNA快速提取[J].科技导报,2013(25):58-60.
[4]田丽波,谷幸幸,商桑,等.苦瓜基因组DNA的提取及ISSR扩增体系的优化[J].中国农学通报,2013,29(4):88-93.
[5]韩玉杰,贾炜珑,王自霞,等.几种提取植物DNA方法的比较[J].山西农业科学,2008,36(7):17-19.
[6]凌莉,李志勇,黄韵,等.食品中植物基因组DNA提取纯化方法研究进展[J].食品科技,2012(5):6-10,15.
[7]张换样,李静,南芝润,等.甘薯DNA的小量快速提取[J].山西农业科学,2009,37(1):12-14.
[8]戴剑,洪德林,张大栋,等.一种快速高效的DNA提取方法研究[J].麦类作物学报,2011(3):437-442.
[9]龚双军,杨立军,刘辉,等.1种小麦白粉病菌DNA基因组的微量简捷提取方法[J].微生物学杂志,2011(1):24-27.
[10]颜松.药用植物基因组DNA提取及铁皮石斛RAPD反应体系的优化[D].重庆:西南交通大学,2009.
[11]张晓祥,王玲,寿路路.一种快速提取小麦基因组DNA的改良CTAB方法[J].中国农学通报,2012,28(36):46-49.
[12]许明,程祖锌,黄志伟,等.一种适于转基因水稻PCR检测的微量DNA快速提取法[J].生物技术通报,2010(3):128-130.
[13]王兰,龙云铭,刘耀光.一种用于PCR的植物基因组DNA快速制备方法[J].分子植物育种,2009,7(2):425-428.
[14]楼巧君,陈亮,罗利军.三种水稻基因组DNA快速提取方法的比较[J].分子植物育种,2005(5):749-752.
[15]任海龙.药用植物大黄种子基因组DNA的提取方法研究[J].山西农业科学,2018,46(3):325-327,349.
[16]刘风路,张金丽,李靖,等.西南地区榧树属植物3种不同总DNA提取方法的比较分析[J].天津农业科学,2018,24(3):1-4.
[17]文静,谢培,孙琛,等.珠子参新鲜和干燥块茎DNA提取方法的比较研究[J].陕西农业科学,2018,64(2):1-3,11.
[18]盛鸥,张玉娥,邓贵明,等.香蕉不同组织的基因组DNA制备方法的比较[J].分子植物育种,2017,15(12):5052-5059.
[19]董永军,王陆军,郝建平,等.玉米干种子基因组DNA提取方法的改进[J].山西农业科学,2017,45(12):1903-1906.
[20]王瑞云,刘笑瑜,王海岗,等.用高基元微卫星标记分析中国糜子遗传多样性[J].中国农业科学,2017,50(20):3848-3870.
[21]闫玖英,马长青,常博,等.改良CTAB法用于苹果果实基因组DNA的提取[J].分子植物育种,2017,15(9):3610-3615.
[22]曾巧英,凌秋平,胡斐,等.5种提取方法对甘蔗汁中DNA提取的效果[J].安徽农业科学,2017,45(25):136-138,152.