Regional mutagenesis using Dissociation in maize

被引:28
作者
Ahern, Kevin R. [1 ]
Deewatthanawong, Prasit [1 ]
Schares, Justin [2 ]
Muszynski, Michael [2 ]
Weeks, Rebecca [2 ]
Vollbrecht, Erik [2 ]
Duvick, Jon [2 ]
Brendel, Volker P. [2 ,3 ]
Brutnell, Thomas P. [1 ]
机构
[1] Cornell Univ, Boyce Thompson Inst Plant Res, Ithaca, NY 14853 USA
[2] Iowa State Univ, Dept Genet Dev & Cell Biol, Ames, IA 50011 USA
[3] Iowa State Univ, Dept Stat, Ames, IA 50011 USA
关键词
Dissociation; Activator; Transposon tagging; Regional mutagenesis; Maize; Functional genomics; ZEA-MAYS L; ACTIVATOR AC; INSERTIONAL MUTAGENESIS; TRANSPOSABLE ELEMENT; DS INSERTION; P-GENE; R-GENE; TRANSPOSITION; EXCISION; ALLELE;
D O I
10.1016/j.ymeth.2009.04.009
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
We describe genetic screens, molecular methods and web resources newly available to utilize Dissociation (Ds) as an insertional mutagen in maize. Over 1700 Ds elements have been distributed throughout the maize genome to serve as donor elements for local or regional mutagenesis. Two genetic screens are described to identify Ds insertions in genes-of-interest (goi). In scheme I, Ds is used to generate insertion alleles when a recessive reference allele is available. A Ds insertion will enable the cloning of the target gene and can be used to create an allelic series. In scheme II, Ds insertions in a goi are identified using a PCR-based screen to identify the rare insertion alleles among a population of testcross progeny. We detail an inverse PCR protocol to rapidly amplify sequences flanking Ds insertion alleles and describe a high-throughput 96-well plate-based DNA extraction method for the recovery of high-quality genomic DNA from seedling tissues. We also describe several web-based tools for browsing, searching and accessing the genetic materials described. The development of these Ds insertion lines promises to greatly accelerate functional genomics studies in maize. (C) 2009 Elsevier Inc. All rights reserved.
引用
收藏
页码:248 / 254
页数:7
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