烟草SCAR标记技术的建立

目的:通过烟草随机扩增多态性DNA(RAPD)标记技术建立烟草特征序列扩增区域(SCAR)标记技术,用于烟草品种鉴定。方法:对12个烟草品种的复烤叶片DNA进行RAPD分析,得到2个RAPD特异片段S1和S2,通过切胶回收,连接pUCm-T载体克隆转化,片段测序,设计特异性引物S1-1/S1-2和S2-1/S2-2,对SCAR-PCR扩增退火温度进行优化。结果:2个RAPD标记成功地转化为稳定快捷的SCAR标记,可将红花大金元和NC102等2个品种从12个烟草品种中快捷准确地鉴别出来。结论:SCAR标记可作为准确稳定的DNA水平的烟草品种鉴定方法,可对种植、复烤和配方品种的烟叶或叶片进行鉴别。

Establishment of Sequence Characteristic Amplified Region Markers of Tobacco

Objective: To develop the sequence characterized amplified region(SCAR) from the random ampli fied polymorphic DNA(RAPD) for identification of tobacco(Nicotiana tabacum L.).Methods: In this study,a total of 12 tobacco species were amplified by 40 randomly selected primers with RAPD method.Two specific and re peatable RAPD markers,S1 and S2,were found.The two specific DNA fragments were cloned and sequenced.Based on the sequencing,two primer pairs,S1-1/S1-2 and S2-1/S2-2,were designed and the SCAR markers for S1 and S2 were obtained.Rresults: The primer pairs S1-1/S1-2 amplify a single 887 bp band in Hongda,K326 and KRK26 but not in other samples,primer pairs S2-1/S2-2 amplify a single 1760 bp band in Hongda and NC102 but not in other samples.The two SCAR markers can accurately and quickly differentiated Hongda and NC102 from other samples.Conclusion: SCAR is a promptly and accurately method can analyses and identify to bacco species at DNA level,and can differentiated tobacco species of tobacco plants,redried tobacco and tobacco of cigarette formula.