Agrobacterium-mediated Genetic Transformation in Cucumber (var. Shital) as Influenced by Explant, Inoculation Time and Co-cultivation Period
Agrobacterium-mediated Genetic Transformation in Cucumber (var. Shital) as Influenced by Explant, Inoculation Time and Co-cultivation Period
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DOI：
10.13189/ujps.2015.030203
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发表时间：
2015
期刊：
影响因子：
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通讯作者：
S. M. Faisal;Muhammad Shahidul Haque;K. M. Nasiruddin
中科院分区：
文献类型：
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作者：
S. M. Faisal;Muhammad Shahidul Haque;K. M. Nasiruddin
为研究黄瓜（Shital var. Shital）非生物抗逆性的遗传转化，利用含有CIPK基因的LBA4404菌株对根癌农杆菌介导的黄瓜叶片、节间和节间愈伤组织进行转化。用组织化学方法检测GUS报告基因在愈伤组织中的转化能力。在愈伤组织中检测到明显的GUS阳性区（蓝色）。这项调查有三个因素。因子A包括叶片、节和节间愈伤组织三种外植体，因子B包括3和5 min两个接种时间，因子C包括24和48 h两个共培养时间。叶片外植体获得的GUS阳性转基因愈伤组织最多（3.30个），节间外植体获得的GUS阳性转基因愈伤组织数量最少（1.79个）。接种时间是影响瘤胃芽胞杆菌转化实验的重要因素。随着接种时间的延长，转化能力增强。愈伤组织在细菌悬浮液中浸泡5 min，愈伤组织成活率较高，为53.68%。GUS阳性愈伤组织培养时间越长（5 min）， GUS阳性愈伤组织的数量和比例越高（分别为3.17%和52.87%），培养时间越短（3 min）， GUS阳性愈伤组织数量和比例越低（分别为2.15%和35.88%）。
To study genetic transformation for abiotic stress resistance in cucumber (var. Shital), leaf, nodal and internodal calli were subjected to Agrobacterium tumefaciens mediated transformation using LBA4404 strain containing CIPK sense gene. Transformation ability was examined by histochemical assay of GUS reporter gene in survived calli. Conspicuous GUS positive (blue colour) region were detected in callus tissue. There were 3 factors in this investigation. Factor A consisted of three types of explants viz. leaf, nodal and internodal callus, factor B consisted of two durations of inoculation time viz. 3 and 5 min and factor C consisted of two co-cultivation periods viz. 24 and 48 hours. The highest GUS positive transgenic callus obtained from leaf explants (3.30) and internodal explants gave the lowest number (1.79) of GUS positive transformants. Inoculation time is an important factor in transformation experiment mediated by A. tumefaciens. Transformation ability was increased with increase of inoculation time. Percentage of survived callus was higher (53.68 %) when the calli were immersed for 5 min in bacterial suspension. Both number and percentage of GUS positive callus were higher (3.17 and 52.87 %, respectively) when they were kept in bacterial suspension for higher time (5 min) and lower (2.15 and 35.88 %, respectively) when calli were soaked in bacterial suspension for minimum time (3 min).