关键词: WRKY转录因子, CaRKNIF2, cDNA, Southern杂交, 基因表达

Abstract:

【Objective】 The aim of the study was to clarify the characterization and expression of one WRKY transcription factor in pepper (Capsicum annuum L. HDA149) during the incompatible interaction between HDA149 and Meloidogyne incognita. 【Method】The combination of RACE and RT-PCR was used to clone the cDNA of a WRKY transcription factor named CaRKNIF2 from pepper inoculated with nematode, and the coding sequences of DNA were amplified with specific primers designed according to the cDNA sequence, copy number of the gene was identified by Southern blot, and the gene expression in various tissue and in different time points after inoculation with nematode were analysed by real-time RT-PCR. 【Result】 The cDNA of CaRKNIF2 (GenBank Accession No: GQ253367) containing an open reading frame of 1 662 bp and encoding 553 amino residues was cloned from pepper. The genome DNA length of CaRKNIF2 was 2 530 bp, and contained 5 introns and 6 exons. CaRKNIF2 was a single copy gene in genome identified by Southern blot. The real-time RT-PCR revealed that the gene expressed tissue-specific with the expression level was the highest in tissue of root tips after inoculation with M. incognita. The expression level of gene CaRKNIF2 in root tips of pepper was increased gradually from 3 to 12 h after inoculation with M. incognita and reached a peak at 12 h. The relative gene expression level at 12 h after nematode inoculation was 3.1 times higher than that of the control. 【Conclusion】 The expression of CaRNIF2 was up-regulated by the nematode inoculation, which implied that the gene might play an important role in the process of incompatible interaction between M. incognita and C. annuum variant HDA149 mediated by resistant gene Me3.

Key words: WRKY transcription factor, CaRKNIF2, cDNA, Southern blot, gene expression