| 熊黎 仝莉 王茂春 周宇荀 李凯 肖君华.特异性逆转录microRNA-505 的高效茎环引物的设计[J].,2015,15(27):5225-5229 |
| 特异性逆转录microRNA-505 的高效茎环引物的设计 |
| Design Specific and Efficient Reverse TranscriptionStem-loop Primer of MicroRNA-505 |
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| DOI： |
| 中文关键词: 茎环引物 miR-505 逆转录 逆转录-聚合酶链式反应（RT-PCR） |
| 英文关键词: Stem-loop primer MiR-505 Reverse transcription RT-PCR |
| 基金项目:中央高校基本科研业务费专项资金(13D110521)；国家自然科学基金项目(31171199) |
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| 中文摘要: |
| 目的：构建一个高效且特异性的逆转录茎环引物，以准确定量检测小鼠的miRNA-505 基因，同时探讨茎环引物的结构对逆
转录效率的影响。方法：规律性地改变通用茎环引物的茎部和环部的碱基结构，再进行不同的组合，利用逆转录-聚合酶链式反应
(Reverse Transcription Polymerase Chain Reaction，RT-PCR)技术，探寻效率最高的茎环引物。结果：当固定茎部结构为14对碱基，
规律性改变环部碱基个数(15，18，21)，经RT-PCR 检测的CT 值无差异，均为28.5；当固定环部结构为21 个碱基，CT 值随茎部碱
基对(8，11，14)规律性地延长而逐渐增大；当环部结构为21 个碱基，茎部结构为8 对碱基时CT 值最小，为26.7。结论：环部结构
对茎环引物的效率无贡献；当环部结构为21 个碱基，茎部结构为8 对碱基时，茎环引物的效率和特异性是最佳的，适用于准确检
测小家鼠的miRNA-505 基因。 |
| 英文摘要: |
| Objective:A high efficient and specific stem-loop primer was designed to detect the miRNA-505 gene accurately in
mice and explore the effect of the stem-loop structure on the efficiency of reverse transcription.Methods:The structures of stempart and
ring part of the general stem-loop primer were changed regularly and put in different combinations to select the optimum stem-loop
primer based on RT-PCR (Reverse Transcription PCR) technology.Results:The results of RT-PCR showed that stem-loop primers,
which was fixed stem structure (14 pairs of bases) and in various loop length (15,18,21bp)had the same CT value (28.5);When the loop
structure was 21 bp, the CT values were gradually increased while the length of stemextended(8,11,14 pairs of bases); The minimumCT
value was obtained when the stem part was 8 pairs of bases and the ring part was 21 bp.Conclusion:The loop structure has no effect on
the reverse transcription. The primer which has 8 pairs of bases stem and 21bp loop has the highest efficiency to the reverse transcription
of miRNA-505 gene in mice. |
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