title: An efficient protocol forAgrobacterium-mediated transformation of the biofuel plantJatropha curcasby optimizing kanamycin concentration and duration of delayed selection.
abstract: Jatropha curcasis considered a potential biodiesel feedstock crop. Currently, the value ofJ. curcasis limited because its seed yield is generally low. Transgenic modification is a promising approach to improve the seed yield ofJ. curcas.AlthoughAgrobacterium-mediated genetic transformation ofJ. curcashas been pursued for several years, the transformation efficiency remains unsatisfying. Therefore, a highly efficient and simpleAgrobacterium-mediated genetic transformation method forJ. curcasshould be developed. We examined and optimized several key factors that affect genetic transformation ofJ. curcasin this study. The results showed that the EHA105 strain was superior to the other threeAgrobacterium tumefaciensstrains for infectingJ. curcascotyledons, and the supplementation of 100 mM acetosyringone slightly increased the transient transformation frequency. Use of the appropriate inoculation method, optimal kanamycin concentration and appropriate duration of delayed selection also improved the efficiency of stable genetic transformation ofJ. curcas. The percentage of β-glucuronidase positiveJ. curcasshoots reached as high as 56.0 %, and 1.70 transformants per explant were obtained with this protocol. Furthermore, we optimized the root-inducing medium to achieve a rooting rate of 84.9 %. Stable integration of the T-DNA into the genomes of putative transgenic lines was confirmed by PCR and Southern blot analysis. Using this improved protocol, a large number of transgenicJ. curcasplantlets can be routinely obtained within approximately 4 months. The detailed information provided here for each step ofJ. curcastransformation should enable successful implementation of this transgenic technology in other laboratories.
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Qiantang
authors.initials: Q
authors.lastname: Fu
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.|College of Life Science and Agriculture, Zhoukou Normal University, Zhoukou, 466001 Henan China.
authors.forename: Chaoqiong
authors.initials: C
authors.lastname: Li
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.|University of Chinese Academy of Sciences, Beijing, 100049 China.
authors.forename: Mingyong
authors.initials: M
authors.lastname: Tang
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Yan-Bin
authors.initials: YB
authors.lastname: Tao
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Bang-Zhen
authors.initials: BZ
authors.lastname: Pan
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Lu
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Longjian
authors.initials: L
authors.lastname: Niu
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Huiying
authors.initials: H
authors.lastname: He
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Xiulan
authors.initials: X
authors.lastname: Wang
authors.affiliation: Key Laboratory of Tropical Plant Resources and Sustainable Use, Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Menglun, 666303 Yunnan China.
authors.forename: Zeng-Fu
authors.identifier: 0000-0001-6045-5865
authors.initials: ZF
authors.lastname: Xu
keywords: Biofuel; Cotyledon; Delayed selection; Kanamycin concentration; Physic nut; Transformation