烤烟CYP82E10基因EMS突变体筛选及其尼古丁转化率分析  

Screening of CYP82E10 mutants from EMS mutant bank of flue-cured tobacco cv. Yunyan 87 and analysis of their nicotine conversion rate

作　　者：李文正[1] 李梅云[1] 吴兴富[1] 王丙武[1] 高玉龙[1] 隋学艺 赵璐 宋中邦[1] LI Wenzheng;LI Meiyun;WU Xingfu;WANG Bingwu;GAO Yulong;SUI Xueyi;ZHAO Lu;SONG Zhongbang(Yunnan Academy of Tobacco Agricultural Sciences,Key Laboratory of Tobacco Biotechnological Breeding,National Tobacco Genetic Engineering Research Center,Kunming 650021,China)

机构地区：[1]云南省烟草农业科学研究院/烟草行业烟草生物技术育种重点实验室/国家烟草基因工程研究中心,昆明650021

出　　处：《中国烟草学报》2018年第5期35-41,共7页Acta Tabacaria Sinica

基　　金：中国烟草总公司云南省公司科技项目(2016YN25);中国烟草总公司科技项目(110201801027(JY-04));国家自然科学基金资助项目(31560080)

摘　　要：烟草尼古丁去甲基酶CYP82E10能够催化尼古丁去甲基化反应,产生致癌物NNN的前体物质。为降低烟叶NNN含量,利用Tilling技术在烤烟云烟87的EMS突变体库中筛选获得烟草尼古丁去甲基酶编码基因CYP82E10的11个突变株,其中M594发生L115F错义突变,M271单株发生P129L错义突变。M4代M594和M271材料的尼古丁转化率分别为对照的45%和38%,M5代分别为对照的34%和45%,说明上述突变能够降低CYP82E10的酶活性及植株转化率。因白肋烟CYP82E10无义突变不影响尼古丁转化率,故烤烟和白肋烟的尼古丁转化机制存在差异。研究获得的低尼古丁转化率突变体可作为育种材料培育低NNN烤烟新品种。Nor-nicotine,precursor of carcinogen N'-nitrosonornicotine(NNN)was derived from nicotine via demethylation process called nicotine conversion which is catalyzed by enzyme nicotine N-demethylase.Minimizing the level of nornicotine in tobacco leaf was believed to be an eff ective way to reduce the harm of cigarette smoke.Two mutant lines were screened out from EMS population of Nicotiana tabacum var.Yunyan87,M594 and M271,with missense mutation in CYP82E10,a gene encoding nicotine N-demethylase,resulted in signifi cant decrease of nicotine conversion rate.In M4 generation,the nicotine conversion rate of M594 and M271 was 45%and 38%of control,respectively.In M5 generation,similar results were observed(34%and 65%of control for M594 and M271 respectively).All these observations were not in line with previous report that the nonsense mutation of CYP82E10 did not change nicotine conversion rate in burley tobacco.An explanation of this discrepancy could be that the fl ue-cured tobacco used in this study did not share same molecular mechanism for the regulation of nornicotine formation with air-cured burley tobacco.Meanwhile,results also showed that both missense mutations could impair the function of CYP82E10.These mutants could be used to develop new tobacco variety with decreased leaf NNN level.

关 键 词：烤烟 NNN CYP82E10 EMS 尼古丁转化率 

分 类 号：S572[农业科学—烟草工业]