GAO Shi-wu, GUO Jin-long, QUE You-xiong, XU Li-ping, YANG Ying-ying, CHEN Ru-kai. Factors affecting transformation efficiency of Agrobacterium tumefaciens EHA105 competent cells[J]. Journal of Tropical Biology, 2012, 3(1): 22-27,31. doi: 10.15886/j.cnki.rdswxb.2012.01.001
| Citation:
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GAO Shi-wu, GUO Jin-long, QUE You-xiong, XU Li-ping, YANG Ying-ying, CHEN Ru-kai. Factors affecting transformation efficiency of Agrobacterium tumefaciens EHA105 competent cells[J]. Journal of Tropical Biology, 2012, 3(1): 22-27,31. doi: 10.15886/j.cnki.rdswxb.2012.01.001
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Factors affecting transformation efficiency of Agrobacterium tumefaciens EHA105 competent cells
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Ministry of Agriculture Key Laboratory of Sugarcane Biology and Genetic Breeding, Fujian Agriculture and Forestry University, Fuzhou 350002, Fuzhou, China
- Received Date: 2011-12-01
- Publish Date:
2012-03-25
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Abstract
Growth of Agrobacterium tumefaciens EHA105,the quality of the competent cells and the heat-shock temperature during transformation are the three main factors affecting the transformation efficiency of A.tumefaciens EHA105.The growth of the strain EHA105 with 48 h continuous culture was observed based on which the growth curve was plotted.At the same time,different concentrations of CaCl2 and different temperatures in heat-shocking during the transformation were tested in preparing the competent cells of EHA105.The number of positive transformants was determined and the factors affecting introduction of exogenous DNA into the strain EHA105 by using the freeze-thaw method were identified.When the OD600 value ranged from 1.0 to 1.5,the strain EHA105 was found at the logarithmic growth phase and had the highest vitality.No significant difference was observed in the transformation rates of competent cells prepared with the CaCl2 at the concentration of between 6 mmol·L-1 and 10 mmol·L-1,but the best transformation rate was 5.322×106·μg-1 at 10 mmol·L-1 CaCl2.No significant difference was found in transformation rates at the heat-shock temperatures from 20 ℃ to 37 ℃ during transformation,but the best transformation rate（5.550×106·μg-1） was achieved at 37 ℃.The results provide a reference for improving the transformation efficiency of A.tumefaciens EHA105 competent cells.
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