杂草稻向转基因水稻基因漂移率的检测技术及验证

摘要：为建立杂草稻向转基因水稻基因漂移后漂移频率的检测技术，将杂草稻和转基因水稻T1c-19按照不同粒数比（1:10-1:100）混合磨样后提取DNA，用杂草稻红皮Rc基因特异性引物进行扩增，观察每种粒数比同时扩增出118bp和104bp的频率，并观察1:60-6:60粒数比时是否可以根据条带颜色深浅判断样品中杂草稻粒数。应用该检测技术，检测了杂草稻WRYY、WRMM向转基因水稻T1c-19的基因漂移率。研究结果表明当杂草稻与转基因水稻粒数比在1:60时，检测阳性频率达到92.5%，但各粒数比下，特异条带颜色无明显差异。因此确定60粒种子为最佳磨样量，但无法根据条带颜色深浅判断磨样中杂交种的粒数。用该方法检测得出两种杂草稻向T1c-19的基因漂移率分别为0.1008%和0.0808%。因此该技术可应用于杂草稻向转基因水稻基因漂移率的检测。

关键词：杂草稻；转基因水稻；基因漂移；检测技术

Detection Technology of Gene Flow Frequency from Weedy Rice to Transgenic Rice and Verification

Abstract：In order to establish a technology for detecting the gene flow frequency from weedy rice to transgenic rice, we mixed weedy rice WRYY and transgenic rice T1c-19 according to different mixed ratios (1:10-1:100), and grinded them into powder to extract DNA. Then we used Rc gene-specific primers to test DNA to observe the frequency of the two bands of 118bp and 104bp in each mixed ratio. Meanwhile we observed whether the number of weedy rice in the sample could be judged according to the color depth of the specific bands in the mixed ratio from 1: 60 to 6: 60. Using this detection technology, we detected gene flow frequency from weedy rice WRYY, WRMM to transgenic rice T1c-19. The results showed that when the mixed ratio of weedy rice and transgenic rice was 1: 60, the positive detection frequency reached 92.5%, but there was no difference in the color depth of the specific bands in different mixed ratios. Therefore, we determined the 60 seeds as the optimal grinding amount, but the number of hybrids in mixed samples could not be judged by the color depth of the bands. The gene flow frequency from weedy rice WRYY and WRMM to transgenic rice T1c-19 was 0.1008% and 0.0808%. The above technology established in this article could be applied to detecting the gene flow frequency from weedy rice to transgenic rice.

Key words: weedy rice；transgenic rice；gene flow；detection technology

摘要3

关键词3

Abstract3

Key words3

引言3

1 材料与方法5

1.1试验材料 5

1.2试验方法 5

1.2.1最佳磨样量的确定5

1.2.2 定量化检测标准的确定6

1.2.3检测杂草稻WRMM和WRYY向转基因水稻T1c-19的基因漂移率6

2 结果与分析6

2.1最佳磨样量的确定6

2.1.1不同粒数比样品总质量及取样量的确定6

2.1.2 DNA浓度的确定6

2.1.3检测不同粒数比中的Rc基因6

2.2定量化检测标准的确定7

2.3杂草稻WRMM和WRYY向转基因水稻T1c-19的基因漂移率检测7

3 讨论9

致谢10

参考文献10

杂草稻向转基因水稻基因漂移率的检测技术及验证

水稻（Oryza sativa L.）是一种重要的粮食作物，世界上近一半的人口以水稻为主食，水稻的丰产和稳产直接关系国家的粮食安全。随着转基因技术的飞速发展，科学家已经培育出了具有抗病虫害、耐盐碱、耐除草剂等具有单一甚至复合性状的转基因水稻[1-4]。我国农业部分别于2009年和2014年两次为华中农业大学培育的转Bt 融合型杀虫蛋白基因 Cry1Ac/Cry1Ab抗虫水稻“华恢1号”和“Bt汕优63”发放了转基因安全证书[5]，这表示转基因水稻向商业化种植的目标又前进了一大步。然而，随着转基因水稻从试验室研究转移到大田种植，也会带来基因漂移的风险。目前关于转基因水稻向杂草稻的基因漂移研究，已经有大量的报道，并建立起了较为完善的研究体系[6-10]。同时有研究表明杂草稻向转基因水稻的基因漂移可能会加速转基因水稻的自身杂草化[11]，但是关于这一方面的研究仍处于起步阶段。本试验旨在探索一种杂草稻向转基因水稻基因漂移率的高效检测技术，并且在实际中进行验证，为转基因水稻生态安全性评估提供科学参考。杂草稻向转基因水稻基因漂移率的检测技术及验证:http://www.chuibin.com/shengwu/lunwen_206223.html