Analysis of Rice Pollen-Specific Promoters

• Main Authors: Yu-Ching Lai, 賴妤青
• Other Authors: 常玉強
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/78989736704831517886

碩士 === 國立臺灣大學 === 農藝學研究所 === 100 === In order to illustrate the functions of pollen-specific genes, putative promoter sequences were isolated from genes in rice pollen with high expression. The putative promoters were constructed with reporter genes, GUS, GFP or a lethal gene (barnase), to explore the specificity of the promoter. First, we screened for the genes with high TPM during the development stages of the pollen based on SBS transcription database. Specific primers were designed to amplify the DNA fragments of 1.5 kb upstream the target genes by PCR, which were named P124 and P128. Also, to characterize the P124 sequences, they were sequentially deleted from the 5’ end to acquire fragments of 1.2, 1.0, 0.9, 0.6, and 0.3 kb. The cloned DNA fragments were then constructed into pCAMBIA1301 and pCAMBIA1303 vectors, and introduced into the rice and Arabidopsis plant via Agrobacteria. Genomic DNA of the transgenic rice was subjected to amplify the flanking sequences by TAIL-PCR. In order to assess the ability of triggering male sterility, P128 was constructed with the barnase gene from Bacillus amyloliquefaciens. The GUS expression was observed mainly in vascular bundle of the anther and petal of the C1301/C1303-P124F transgenic rice as well as in vascular bundle of the anther and pollen of the transgenic Arabidopsis. Additionally, GUS was significantly expressed in the anther and pollen of the transgenic rice containing C1301-P128. The sequential deletion analysis of P128 contributes to further research in male sterility from inactive pollens.