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Acta Agron Sin ›› 2011, Vol. 37 ›› Issue (02): 374-379.doi: 10.3724/SP.J.1006.2011.00374

• RESEARCH NOTES • Previous Articles    

Cloning of Medicago sativa Phychrome B cDNA and Establishment of Its RNA Interference Expression Vector

ZHU Jian-Ming,YAN Xue-Bing,SHI Ying-Hua,WANG Cheng-Zhang*   

  1. Engineering College of Animal Husbandry and Veterinary Sciences, Henan Agricultural University, Zhengzhou 450002, China
  • Received:2010-05-28 Revised:2010-09-22 Online:2011-02-12 Published:2010-12-15
  • Contact: 王成章, E-mail:wangchengzhang@263.net

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Abstract: The short day is one of the main factors affecting alfalfa fall dormancy, which is called as the photoperiodic effect. Study on the relationship between the main photoreceptors-PhyB gene and alfalfa fall dormancy may reveal the regulation mechanism of alfalfa fall dormancy radically, and provide a scientific reference for the application of alfalfa varieties differing in fall dormancy in forage production. The objective of this study was to establish an RNAi expression vector harboring PhyB gene of Medicago sativa. Two pairs of specific primers containing different enzyme sites were designed on the basis of PhyB gene sequence of alfalfa variety “Vernal” (GenBank accession number: GQ379903.1). With the template of total RNA, positive sense strand and antisense strand were amplified by RT-PCR and cloned into pGEM-T Easy vector to obtain recombinant vectors pGEMB-1 and pGEMB-2, then based on the intermediate vector pHANNIBAL and the plant binary expression vector pART27, we constructed the RNAi expression vector pART27-RNAi containing a hairpin structure by many times of enzyme digestion and connection. The results provide a foundation for further studying the relationship between alfalfa dormancy and PhyB.

Key words: Medicago sativa, Fall dormancy, Photoreceptor, Phytochrome, RNA interference

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