Identification Of Partner Genes Interact With AtMT-A70B By Yeast-2-Hybrid System

The pre-mRNAs undergo a number of post-transcriptional modification steps prior to translocation to the cytoplasm where they are translated into proteins. These steps include capping polyadenylation and pre-mRNA splicing. In addition to these obvious processing steps, post-transcriptional modification of certain nucleosides also occurs~[1]. N~6-Methyladenosine has been proposed to play a role in mRNA processing and transport. In recent years, research focusing on RNA molecular biology has shown that this post-transcriptional event plays important roles in the overall regulation of gene expression. In arabidopsis the genes responsible for N~6-Methyladenosine Methyltransferase were proved essential for embryo genesis by Dr Rupert .G. Fray's group.In order to find the other partners in Methyltransferation, this thesis mainly describe the cloning of MB*(AtMT-A70B) from wild type arobidopsis using specific primers (PCR and RT-PCR) and then using the MB* as bait to screen Arabidopsis developing anther yeast two-hybrid library. Colonies Bin1/2 Bin1/14 Bin1/20 Binl3/1 and Bin15/7 have been proved positive reactions. After these colonies have been sequenced, NCB-blast analysis of these interactions proves Bin15/7 Binl/14 Binl/20 are protein At5g49020 and Bin13/1 Bin1/2 are protein At3g61690. At5g49020's function in Arabidopsis is N-methyltransferase protein. Both of N-methyltransferase and At3g61690 take part in Methyltransferation is found firstly. These results are very important for further understanding of the additional proteins required for mRNA methylating activity. During the procedure, different molecular biology techniques were involved such as PCR RT-PCR and Yeast-two-hybrid system.