轉殖順義ACC合成酶基因延緩苦瓜果實後熟之研究

Studies on Delaying Fruit Ripening by Transforming Sense ACC Synthase Gene into Momordica charantia L.

林生華(Shung-Hua Lin) ； 杜宜殷(Yi-Yin Do)

《臺灣園藝》 54卷2期 (2008/06) Pp. 115-125

https://doi.org/10.6964/JTSHS.200806.0115

摘要

本研究運用苦瓜花粉電穿孔法，將含順義苦瓜ACC合成酶(ACC synthase, ACS)cDNA之質體DNA導入苦瓜基因組中後，經以100 µg•mL^(-1)抗生素Kanamycin篩選轉殖植株，存活的轉殖植株繼續進行GUS活性組織化學染色分析，進一步抽取呈現藍色正反應的轉殖植株之基因組DNA，以ACS cDNA、GUS及NPT II基因片段等3個探針，進行基因組DNA南方氏雜交分析。花粉電穿孔法雖可取得具轉殖基因之植株，但僅有部份外來轉殖基因整合入苦瓜基因組內。轉殖植株(GS2063)扦插繁殖，結果成熟後，測定其乙烯生成。苦瓜順義ACS基因轉殖株之果實乙烯生成高峰，較非轉殖對照果實遲2天出現，且高峰值低16%。轉殖順義ACS基因有可能延緩苦瓜果實後熟。

關鍵字

基因默化 ； 花粉電穿孔法 ； 南方氏雜交分析 ； 乙烯生合成

並列摘要

A plasmid containing sense ACC synthase (ACS) gene was introduced to bitter gourd (Momordica charantia L.) genome by pollen electroporation method. Some putative transgenic seeds germinated and were selected by spraying 100 µg•mL^(-1) Kanamycin. Leaves of the survived seedlings were analyzed by histochemical staining of β-glucuronidase (GUS) activity. Genomic DNA of putative transgenic plants was extracted individually and subjected to Southern analysis using ACS cDNA, GUS gene and NPTII gene as probes. Only partial fragment of transgene was integrated into the genome of bitter gourd via pollen electroporation. Transformant line GS2063 was chosen for further propagation by cutting. Mature fruits of the transgenic bitter gourd were harvested and the rate of ethylene production of each fruit was measured daily. When kept at 25℃, fruits harvested from sense ACS gene transformants took 2 days longer to reach ethylene production peak and 16% lower peak value than fruits from selfing progeny of untransformed plants. Transformation of sense ACC synthase gene may generate a slower ripening bitter gourd.

並列關鍵字

gene silencing ； pollen electroporation ； Southern hybridization ； ethylene biosynthesis