Cloning And Functional Analysis Of DnWRKY11Transcription Factor

Dendrobium species are epiphytic plants of Orchidaceae. Many wild species of Dendrobium are precious ornamental plants and traditional precious Chinese medical herbals. The family of WRKY transcription factors is very important. It plays important roles in plant growth, development, senescence and adapting to biotic or abiotic stress. Study the function of WRKY transcription factors in Dendrobium species is important for providing theoretical and application value. In the present study, we identified a Ⅱ WRKY gene from Dendrobium nobile, and explored its function by the methods of bioinformatic analysis, tissue-specific expression analysis and over-expression tobacco. The main results are as follows.(1) In the present study, parts of cDNA sequences were chosen from our previous laboratory cDNA library. One of the full length of the WRKY transcription factor cDNA was obtained by the RACE technology and nominated DnWRKYll. Bioinformatic analysis showed that its full-length sequence is1135bp, and contains47bp of the51non-coding region and167bp of the3’non-coding region. It encodes306amino acids, which containing a WRKY structure domain and considered to be the zinc finger C2H2type.(2) By analyzing the tissue-specific expression of DnWRKY11in Dendrobium nobile with the methods of real-time qPCR, we found the DnWRKY11is highest expression in leaves, followed by roots, and the lowest is in the stem of Dendrobium nobile. The expression in leaves is4.5times higher than that in the root.(3) The pSN1301-DnWRKY11was built and transformed tobacco by Agrobacterium transformation. DnWRKY11overexpression plants were obtained. Their seeds and seedlings response to drought (mannitol) and salt (NaCl) stress was studied. In the present results, compared to wild-type tobacco, transgenic tobacco seeds and seedlings showed more resisitence to drought and salt stress.The germination rate of transgenic tobacco seeds was higher than wild type. The root length of transgenic tobacco seedlings was longer than wild type. (4) The wild type had higher malonidialdehyde content than the transgenic tobacco line in drought stress. What’s more, the activity of three enzymes that related to oxidative stress in the transgenic tobacco was higher than that in the wild type. The transgenic tobacco was higher oxidative resistance. It suggested that DnWRKY11transcription factors played a positive role in this process. By analyzing the expression of drought gene NtERD in wild-type tobacco and transgenic tobacco seedlings at200mM mannitol with the methods of real-time qPCR, we found the expression of NtERD in over-expression tobacco was uper than wild-type tobacco. The expression in T1-22transgenic line was5.4times higher than that in wild-type tobacco.These results show that DnWRKY11transcription factor was involved in the reaction of plant resistance to abiotic stresses, drought and salt stress reaction. It is a positive regulatory factor and plays an active role.