盐芥miRNAs耐盐表达研究  

Studies on Expression of miRNAs in Thellungiella under Salt Stress

作　　者：刘冉[1] 周立敬[1] 周宜君[1] 高飞[1] 马亭亭[1] 隋欣[1] 刘楠[1] 

机构地区：[1]中央民族大学生命与环境科学学院,北京100081

出　　处：《西北植物学报》2012年第4期665-669,共5页Acta Botanica Boreali-Occidentalia Sinica

基　　金：国家自然科学基金(31070361);中央高校基本科研业务费专项(0910KYZY43;1112KYQN31);中央民族大学"985工程"项目(MUC98504-14);国家民委科研项目(10ZY01)

摘　　要：以盐生植物盐芥为实验材料,选择经过Solexa测序筛选的盐芥tsa-miR172a和tsa-miR398b为目标基因,采用茎环的反转录PCR(stem-loop RT-PCR)方法分析其在盐芥根中的耐盐表达模式,以探讨盐芥miRNAs的stem-loop RT-PCR验证体系。结果显示,经过300mmol.L-1 NaCl处理72h后,与对照相比,盐芥根中的tsa-miR172a上调表达,tsa-miR398b下调表达。用stem-loop RT-PCR方法进行tsa-miR172a和tsa-miR398b扩增,对其电泳图进行光密度分析结果显示,盐胁迫处理与对照的比值分别为1.8和0.55,Solexa测序结果分别为2.00和0.44,说明两种方法所得结果基本一致。表明该研究建立了盐芥miRNAs的stem-loop RT-PCR验证体系:每个miRNA设计3个引物(miRNA stem-loop引物、miRNA正向引物和miRNA通用反向引物);扩增条件为94℃2min,94℃15s,55℃45s,23个循环。The expressions of tsa-miR172a and tsa-miR398b in roots of Thellungiella halophila （T. salsug- inea） ,which were screened via Solexa sequencing,were studied by stem-loop RT-PCR methodology. The results showed that tsa-miR 172a was up-expressing and tsa-miR 398b was down-expressing under the treat- ment of NaC1 （300 mmol · L^-1, 72 h）. With the stem-loop RT-PCR,tsa-miR172a and tsa-miR398b were amplified and the optical densities of electrophores were analyzed specifically. The ratios of NaC1 stress to control were 1.8 and 0.55 ,which were roughly consistent with the results 2.00 and 0.44 obtained by Sol- exa. Our study established an effective stem-loop RT-PCR system：three primers,namely miRNA stem-loop primer,miRNA forward primer and miRNA universal reverse primer,were designed for each miRNA. The reactions were incubated at 94℃ for 2 rain,followed by 23 cycles of 94℃ for 15 s and 55℃ for 45 s. The stem-loop RT-PCR system to identify miRNAs in Thellungiella was discussed.

关 键 词：盐芥 MIRNAS 茎环的反转录PCR 盐胁迫 

分 类 号：Q789[生物学—分子生物学]