Construction and analysis of transgenic Arabidopsis with AtMAPRs gene silencing vectors

• Main Authors: Shang-Hsuan Yeh, 葉尚烜
• Other Authors: 楊健志
• Format: Others
• Language: zh-TW
• Published: 2006
• Online Access: http://ndltd.ncl.edu.tw/handle/65194948824178974967

碩士 === 國立臺灣大學 === 微生物與生化學研究所 === 94 === Four AtMAPRs (AtMAPR2, AtMAPR3, AtMPAR4, and AtMAPR5) in Arabidopsis sharing 30-40% similarity with porcine MAPR (membrane associated progesterone receptor component 1) have been identified. AtMAPR5 (putative membrane steroid binding protein, MSBP1) is proposed as a negative regulator of cell elongation (Yang et al., 2005). However, the functions of the other AtMAPRs are still unclear. By Agrobacterium-mediated transformation, we attempted to establish two transgenic plants where AtMAPR4 gene was either knocked-down by RNAi technology or overexpressed. Transgenic plants overexpressing AtMAPR4 showed that their bolting time was earlier than wild-type, whereas transgenic plants with knocked-down AtMAPR4 showed that they delayed flowering. We also found that AtMAPR4 was expressed higher in flowers by analyzing the tissue-specific expression in 35-day-old wild-type plants previously. It suggested that AtMAPR4 might be involved in the flowering. The second part of this research followed previous yeast two-hybrid assay experiment where AtMAPR5 might interact with RING-type ubiquitin ligase (At3g58030). Another RING-type ubiquitin ligase (At2g42030) shares high similarity with At3g58030. RING-type ubiquitin ligase could recognize target protein by specific domain. These two RING-type ubiquitin ligase were cloned and heterologously expressed in E.coli. They will be used in further pull-down experiment and ubiquitination assay.