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Acta Agron Sin ›› 2015, Vol. 41 ›› Issue (12): 1810-1818.doi: 10.3724/SP.J.1006.2015.01810

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Cloning and Characterization of a Novel Gene Encoding Proline-Rich Protein in Sesame

LÜ Gao-Qiang,WU Xiang-Yang,WANG Xin-Yu*   

  1. College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, China
  • Received:2015-04-13 Revised:2015-07-20 Online:2015-12-12 Published:2015-08-28
  • Contact: 王心宇, E-mail: xywang@njau.edu.cn
  • Supported by:

    This research was supported by the National Basic Research Program of China (973 Program) (2011CB109300).

Abstract:

Bacterial wilt of sesame is a major threat in sesame production in south China, resulting seriously in yield and quality losses. The disease is caused by bacterial pathogen Ralstonia solanacearum. This study profiled the gene expression of sesame inoculated with Ralstonia solanacearum by using fifty random primers. A gene (fragment) was found to be drastically down regulated by the pathogen. The gene fragment was cloned and sequenced. Using the sequence as queries, the sesame genome database (http://www.ncbi.nlm.nih.gov/) was searched and the corresponding DNA sequence containing a complete ORF was obtained. The full-length of the gene shows that its encoding region is 1458 bp, encoding a putative protein of 486 amino acids. The protein is rich in proline on its N-terminus, and has several repeat sequences (motifs) rich in proline, suggesting that it belongs to proline-rich protein (PRP) family. The protein was named as SiPRP (Sesaumu indicunm Proline-rich Protein). The encoding region of SiPRP was further amplified in sesame cDNAs, sequencing analysis demonstrated that it has the same sequence with the predicted one. Blast analysis revealed that the protein has the lower homology with other plant PRPs, and has new types of proline-rich motifs, suggesting that SiPRP is a new member in PRP family. Semi quantitative RT-PCR and qPCR with newly designed gene-specific primers verified that SiPRP expression was drastically down regulated upon pathogen infection. Previous studies showed that most plant PRPs were located on plant cell wall, however, transient expression in onion epidermal cells showed that SiPRP-YFP fusion protein was located on cell membrane, with a bit secreted outside the cell. Transient expression in tobacco cells revealed that SiPRP protein might be located on special structures of the membrane. SiPRP protein identified in this study may play pivotal roles in Ralstonia solanacearum-sesame interactions.

Key words: Sesame (Sesaumu indicunm), Ralstonia solanacearum, Proline-rich protein, Induced expression, Subcellular localization





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