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標題: 鐵炮百合兩個嶄新的花藥專一基因與其蛋白質之特性分析
Characterization of Two Novel Anther-specific Genes and Their Encoding Proteins in Lilium longiflorum
作者: 楊正守
Yang, Cheng-Shou
關鍵字: Lilium longiforum;花藥;anther;tapetum;exine;绒氈層;花粉外壁
出版社: 生物科技學研究所
引用: 曾稚鐙。2006鐵炮百合花藥發與早期基因之選殖與特性分析。國立中興大學碩士論文。 葉芳凌。2008鐵炮百合兩绒氈層基因LLA-1271和LLA-115之特性分析和表現。國立中興大學碩士論文。 Aarts MG, Hodge R, Kalantidis K, Florack D, Wilson ZA, Mulligan BJ, Stiekema WJ, Scott R, Pereira A (1997) The Arabidopsis MALE STERILITY 2 protein shares similarity with reductases in elongation/condensation complexes. Plant J 12: 615-623 Achard P, Baghour M, Chapple A, Hedden P, Van Der Straeten D, Genschik P, Moritz T, Harberd NP (2007) The plant stress hormone ethylene controls floral transition via DELLA-dependent regulation of floral meristem-identity genes. Proc Natl Acad Sci U S A 104: 6484-6489 Ariizumi T, Hatakeyama K, Hinata K, Inatsugi R, Nishida I, Sato S, Kato T, Tabata S, Toriyama K (2004) Disruption of the novel plant protein NEF1 affects lipid accumulation in the plastids of the tapetum and exine formation of pollen, resulting in male sterility in Arabidopsis thaliana. 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LLA-142和LLA-1271是利用抑制扣除雜合法(suppression subtractive hybridization)從鐵炮百合小孢子發育時期的花藥cDNA集合庫中挑出的cDNA片段。LLA-142和LLA-1271利用5′與3′-RACE得到全長的序列。LLA-142全長為379 bp,可轉譯出56個胺基酸,估計蛋白質分子量為6.2 kDa。LLA-1271 cDNA可發現兩條類似的序列,LLA-1271a全長為970 bp可轉譯出225個胺基酸,而LLA-1271b全長為 879 bp可轉譯出226個胺基酸,其差異為LLA-1271a在ORF少了3 bp,而LLA-1271b在3′-UTR缺少了一段76 bp的序列。LLA-1271扣除N端26個胺基酸訊息胜肽,估計成熟蛋白質分子量為21 kDa,整個蛋白質為八個高度保留的重複序列所涵蓋。LLA-142和LLA-1271經由北方墨漬法分析,顯示出兩基因都為小孢子時期花藥專一性的基因,在小孢子及花藥壁都有表現。進一步利用原位雜合(in situ hybridization)的實驗確定LLA-142和LLA-1271在花藥的绒氈層表現,在花藥發育的早期(花苞2 cm)並無偵測到兩基因的訊號,3-4 cm花苞時在花藥的绒氈層有明顯的訊號,5 cm花苞時绒氈層的兩基因訊號減弱,顯示出兩基因表現的強弱隨著绒氈層發育同步消長。LLA-1271 mRNAs的表現受內生和外加的激勃素誘導,而LLA-142不受外加的激勃素所調節。利用激勃素抑制劑uniconazole和乙烯抑制劑2,5-norbornadiene處理,顯示LLA-1271受激勃素和乙烯的調控,並且乙烯和激勃素相互影響LLA-1271的表現。利用西方墨漬法分析,發現LLA-142和LLA-1271為小孢子時期花藥專一性的蛋白質。利用花藥分層分析顯示在處理過0.5%或2% Triton X-100的小孢子之壁蛋白中,可偵測到LLA-1271蛋白訊號。LLA-1271累積在小孢子發生有絲分裂之前的外壁上,推測其蛋白質可能參與小孢子外壁早期的發育。利用TAIL-PCR找到LLA-142基因的啟動子序列966 bp,其中包含了TATA-box、CAAT-box、七個花粉專一調控區(AGAAA、TCAC)和荷爾蒙相關的調控區。
其他識別: U0005-2107200815402100
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