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A Ligation-Independent Cloning Tobacco Rattle Virus Vector for High-Throughput Virus-Induced Gene Silencing Identifies Roles for NbMADS4-1 and -2 in Floral Development

Yiyu Dong, Tessa M. Burch-Smith, Yule Liu, Padmavathi Mamillapalli and Savithramma P. Dinesh-Kumar
Plant Physiology
Vol. 145, No. 4 (Dec., 2007), pp. 1161-1170
Stable URL: http://www.jstor.org/stable/40065758
Page Count: 10
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Since scans are not currently available to screen readers, please contact JSTOR User Support for access. We'll provide a PDF copy for your screen reader.
A Ligation-Independent Cloning Tobacco Rattle Virus Vector for High-Throughput Virus-Induced Gene Silencing Identifies Roles for NbMADS4-1 and -2 in Floral Development
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Abstract

Virus-induced gene silencing (VIGS) is a widely used, powerful technique for reverse genetics. VIGS vectors derived from the Tobacco rattle virus (TRV) are among the most popular for VIGS. We have developed a TRV RNA2 vector that allows the insertion of gene silencing fragments by ligation-independent cloning (LIC). This new vector has several advantages over previous vectors, particularly for applications involving the analysis of large numbers of sequences, since TRV-LIC vectors containing the desired insert are obtained with 100% efficiency. Importantly, this vector allows the high-throughput cloning of silencing fragments without the use of costly enzymes required for recombination, as is the case with GATEWAY-based vectors. We generated a collection of silencing vectors based on 400 tomato (Solanum lycopersicum) expressed sequence tags in this TRV-LIC background. We have used this vector to identify roles for SIMADS1 and its Nicotiana benthamiana homologs, NbMADS4-1 and -2 in flowering. We find that NbMADS4-1 and -2 act nonredundantly in floral development and silencing of either gene results in loss of organ identity. This TRV-LIC vector should be a valuable resource to the plant community.

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