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Acta Agron Sin ›› 2015, Vol. 41 ›› Issue (07): 1056-1063.doi: 10.3724/SP.J.1006.2015.01056


Screening of AhCaM-Interactive Proteins in Peanuts Using Yeast Two Hybrid System

YANG Sha1,LI Yan1,GUO Feng1,ZHANG Jia-Lei1,MENG Jing-Jing1,LI Meng2,WAN Shu-Bo1,*,LI Xin-Guo1,*   

  1. 1 Biotechnology Research Center of Shandong Academy of Agricultural Sciences / Shandong Provincial key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Ji’nan 250100, China; 2 Human Resources Department of Shandong Academy of Agricultural Sciences Ji’nan 250100, China
  • Received:2014-11-06 Revised:2015-05-04 Online:2015-07-12 Published:2015-05-15
  • Contact: 万书波, E-mail: wansb@saas.ac.cn; 李新国, E-mail: lixinguo@tom.com E-mail:yangsha0904@126.com


Calmodulin is the most important known receptor of intracellular Ca2+, which is involved in regulation of many physiological activities. To further study the mechanism of CaM, we isolated and identified proteins that interact with AhCaM by yeast two-hybrid system. The pGBKT7-AhCaM bait vector without toxicity or auto-activation was firstly constructed with the peanut CaM gene. Then, the total RNA of peanut leaf was extracted and the mRNA was isolated, purified to be used as the templates to synthesize ds-cDNA by SMART technology. Ds-cDNA was amplified by long distance PCR. At last, the CaM interaction proteins were screened through co-transformation with bait vector and cDNA prey library. In the peanut cDNA library, five proteins interacting with AhCaM were identified. Among them, NAD kinase was a well-known protein to interact with CaM protein, and ubiquitin could interact with AhCaM, playing a role in development and stress resistance. The significant functional correlation between AhCaM and its interacting proteins peanut will help to elucidate the possible mechanisms of AhCaM in improving the tolerance of transgenic plants.

Key words: cDNA library of peanut leaf, AhCaM, Ubiquitin, Yeast two hybrid system, Protein interaction

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