Stable chloroplast transformation of immature scutella and inflorescences in wheat (Triticum aestivum L.)
- Cuiju Cui†,
- Fei Song†,
- Yi Tan,
- Xuan Zhou,
- Wen Zhao,
- Fengyun Ma,
- Yunyi Liu,
- Javeed Hussain,
- Yuesheng Wang,
- Guangxiao Yang and
- Guangyuan He*
- China-UK HUST-RRes Genetic Engineering and Genomics Joint Laboratory, The Genetic Engineering International Cooperation Base of Ministry of Science and Technology, the Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science & Technology, Wuhan 430074, China
- *↵Correspondence address. Tel: +86-27-87792271; Fax: +86-27-87792272; E-mail: hegy{at}hust.edu.cn
-
↵† These authors contributed equally to this work.
- Received September 25, 2010.
- Accepted December 27, 2010.
Abstract
Chloroplast transformation in wheat was achieved by bombardment of scutella from immature embryos and immature inflorescences, respectively. A wheat chloroplast site-specific expression vector, pBAGNRK, was constructed by placing an expression cassette containing neomycin phosphotransferase II (nptII) and green fluorescent protein (gfp) as selection and reporter genes, respectively, in the intergenic spacer between atpB and rbcL of wheat chloroplast genome. Integration of gfp gene in the plastome was identified by polymerase chain reaction (PCR) analysis and Southern blotting using gfp gene as a probe. Expression of GFP protein was examined by western blot. Three positive transformants were obtained and the Southern blot of partial fragment of atpB and rbcL (targeting site) probes verified that one of them was homoplasmic. Stable expression of GFP fluorescence was confirmed by confocal microscopy in the leaf tissues from T1 progeny seedlings. PCR analysis of gfp gene also confirmed the inheritance of transgene in the T1 progeny. These results strengthen the feasibility of wheat chloroplast transformation and also give a novel method for the introduction of important agronomic traits in wheat through chloroplast transformation.
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- © The Author 2011. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.
