Gene identification with sequenced T‐DNA tags generated by transformation of Arabidopsis cell suspension

Gene identification with sequenced T‐DNA tags generated by transformation of Arabidopsis cell... A protocol for establishment and high‐frequency Agrobacterium‐mediated transformation of morphogenic Arabidopsis cell suspensions was developed to facilitate saturation mutagenesis and identification of plant genes by sequenced T‐DNA tags. Thirty‐two self‐circularized T‐DNA tagged chromosomal loci were isolated from 21 transgenic plants by plasmid rescue and long‐range inverse polymerase chain reaction (LR‐iPCR). By bidirectional sequencing of the ends of T‐DNA‐linked plant DNA segments, nine T‐DNA inserts were thus localized in genes coding for the Arabidopsis ASK1 kinase, cyclin 3b, J‐domain protein, farnesyl diphosphate synthase, ORF02, an unknown EST, and homologues of a copper amine oxidase, a peripheral Golgi protein and a maize pollen‐specific transcript. In addition, 16 genes were identified in the vicinity of sequenced T‐DNA tags illustrating the efficiency of genome analysis by insertional mutagenesis. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png The Plant Journal Wiley

Gene identification with sequenced T‐DNA tags generated by transformation of Arabidopsis cell suspension

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Publisher
Wiley
Copyright
Copyright © 1998 Wiley Subscription Services, Inc., A Wiley Company
ISSN
0960-7412
eISSN
1365-313X
DOI
10.1046/j.1365-313X.1998.00059.x
Publisher site
See Article on Publisher Site

Abstract

A protocol for establishment and high‐frequency Agrobacterium‐mediated transformation of morphogenic Arabidopsis cell suspensions was developed to facilitate saturation mutagenesis and identification of plant genes by sequenced T‐DNA tags. Thirty‐two self‐circularized T‐DNA tagged chromosomal loci were isolated from 21 transgenic plants by plasmid rescue and long‐range inverse polymerase chain reaction (LR‐iPCR). By bidirectional sequencing of the ends of T‐DNA‐linked plant DNA segments, nine T‐DNA inserts were thus localized in genes coding for the Arabidopsis ASK1 kinase, cyclin 3b, J‐domain protein, farnesyl diphosphate synthase, ORF02, an unknown EST, and homologues of a copper amine oxidase, a peripheral Golgi protein and a maize pollen‐specific transcript. In addition, 16 genes were identified in the vicinity of sequenced T‐DNA tags illustrating the efficiency of genome analysis by insertional mutagenesis.

Journal

The Plant JournalWiley

Published: Mar 1, 1998

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