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Gene identification with sequenced T‐DNA tags generated by transformation of Arabidopsis cell suspension
Author(s) -
Mathur Jaideep,
Szabados László,
Schaefer Sabine,
Grunenberg Britta,
Lossow Andrea,
JonasStraube Esther,
Schell Jeff,
Koncz Csaba,
KonczKálmán Zsuzsanna
Publication year - 1998
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1046/j.1365-313x.1998.00059.x
Subject(s) - biology , arabidopsis , gene , insertional mutagenesis , agrobacterium , transformation (genetics) , genetics , transfer dna , microbiology and biotechnology , genome , mutant
Summary 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.