Plant Diversity ›› 2014, Vol. 36 ›› Issue (05): 622-628.doi: 10.7677/ynzwyj201413225

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Cloning of ACC Oxidase Gene from Yali Pear and Tranformation of Its Antisense Expression Vector with Agrobacteriummediated Method

QI Jing-1, DONG Zhen-2, ZHANG Yu-Xing-3   

  1. 1 College of Biology Science and Engineering, Hebei University of Economics and Business, Shijiazhuang 050061, China; 2 Hebei Women′s Vocational College, Shijiazhuang 050091, China; 3 College of Horticulture, Agriculture University of Hebei, Baoding 071001, China
  • Received:2013-11-20 Online:2014-09-25 Published:2014-02-20
  • Supported by:

    河北省科技计划资助项目 (11220103D9);河北省高等学校科学技术研究指导项目 (Z2012065)

Abstract:

In this study, a partial ACC oxidase (ACO) genelike cDNA sequence was obtained through homologybased cloning from Yali (Pyrus bretschneideri cv. ‘Yali’) plant. Primers were designed according to the highly conserved regions of published ACO gene sequences, and RTPCR cloning was conducted by using Yali fruit cDNA. The obtained ACOlike cDNA fragment contains 831 base pairs which encodes 276 predicted amino acid residues, and shares no less than 94% nucleotide sequence identity with all published ACO genes. We further inversely inser ted the ACOlike cDNA fragment into pBI121 expression vector, and transformed it into tissue cultured Yali plants by using Agrobacterium LBA4404. Finally, 4 independent transgenic lines harboring the antisense ACOlike fragment were obtained and validated by PCR analysis. Southern blotting assay revealed 3 transgenic lines containing single copy of the foreign gene, and 1 line with 2 insertion copies.

Key words: Yali pear, ACC oxidase, RT-PCR, Antisense expression vector, Genetic transformation

CLC Number: 

  • Q 784