Czech J. Genet. Plant Breed., 2013, 49(2):51-57 | DOI: 10.17221/217/2012-CJGPB

Integration and characterization of T-DNA insertion in upland cottonOriginal Paper

Xiaojie YANG^1,2, Fuguang LI¹, Xueyan ZHANG¹, Kun LIU¹, Qianhua WANG¹, Chaojun ZHANG¹, Chuanliang LIU¹, Wei ZHU³, Guofang SHAN¹, Chee-Kok CHIN⁴, Weiping FANG²

¹ State Key Laboratory of Cotton Biology, Institute of Cotton Research, Chinese Academy of Agriculture Sciences (CAAS), Anyang, Henan, P.R. China

² Economic Crop Research Institute, Henan Academy of Agricultural Sciences, Zhengzhou, P.R. China

³ Agronomy College of Henan Agricultural University, Zhengzhou, P.R. China

⁴ Department of Plant Science, Rutgers University, New Jersey, USA

Copy numbers were evaluated by real-time quantitative PCR, and 149 junctions of T-DNA were isolated by thermal asymmetric interlaced PCR from 92 independent transgenic cotton lines transformed by Agrobacterium tumefaciens strain LBA4404. Real-time quantitative PCR results showed that 46% had integration of one or two T-DNA copies, 54% had three or more copies. Among 63 amplified products at LB junctions, 51% showed co-transformation of the vector backbone, 30% retained a portion of LB ranging from 3 to 23 bp, and 19% showed deletions ranging from 1 to 148 bp from the LB inner end. In contrast, all of the cleavage sites were located in the inner region of RB. The distribution of T-DNA insertions in upland cotton genome included coding sequences, transposons, plastid-derived sequences and microsatellites.

Keywords: cotton (Gossypium hirsutum L.); deletion of border; genetic transformation; transgene copy; vector integration

Published: June 30, 2013  Show citation