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Use of a herbicide or lysine and threonine for non-antibiotic selection of transgenic chickpea

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Title Use of a herbicide or lysine and threonine for non-antibiotic selection of transgenic chickpea
 
Creator Tewari-Singh, N.
Sen, Jayanti
Kiesecker, H.
Reddy, V. S.
Jacobsen, H.J.
Guha-Mukherjee, S.
 
Subject Chickpea (Cicer arietinum L.)
Aspartate kinase
Lysine plus threonine
Phosphinothricin
Transformation
 
Description A desensitized aspartate kinase (AK) gene has
been developed as a non-antibiotic selection marker for
use in the production of transgenic chickpea (Cicer
arietinum L.). Transgenic shoots regenerated from embryo explants bombarded with the desensitized AK gene
were selected on media containing two amino acids,
lysine and threonine (LT). Approximately 15% of the
putative transgenic shoots of vars. P-362 and P-1042
survived after 4 weeks of growth on MSB5 medium (MS
mineral salts and B5 vitamins) containing 2 M thidiazuron (TDZ) and 2 mM lysine and 2 mM threonine.
These shoots were subsequently grown on MSB5 medium
supplemented with 2 M TDZ and 5 mM lysine and 5 mM threonine, and nearly 1% continued to grow after
16 weeks of selection. A phosphinothricin (PPT) selection
system for Agrobacterium-mediated chickpea transformation was also developed. Three varieties of chickpea,
P-362, P-1042 and P-1043, were successfully used for
Agrobacterium transformation. Following Agrobacterium
infection, 3-8% of the regenerated shoots remained green and continued to grow on MSB5 medium supplemented
with 2.5 mg l-1 PPT. Increasing the concentrations of
PPT to 15 mg l-1 reduced transgenic shoot production in
P-362, P-1042 and P-1043 to 0.7%, 1.2% and 1.1%,
respectively. Selected putatively transformed shoots of all
three varieties were rooted and grown to maturity.
Southern hybridization analysis revealed single as well
as multiple integration of genes in selected transgenic
lines. The level of AK activity detected in LT-selected
plants was higher than that detected in the non-transformed control.
This work was supported by grants from the
Department of Biotechnology, Government of India to SGM.
 
Date 2013-10-29T10:42:11Z
2013-10-29T10:42:11Z
2004
23 September 2003
 
Type Article
 
Identifier Plant Cell Reports, 22: 576-583
http://hdl.handle.net/123456789/39
 
Language en
 
Publisher Springer