Targeted parallel sequencing of large genetically-defined genomic regions for identifying mutations in Arabidopsis

被引:11
|
作者
Liu, Kun-hsiang [1 ,2 ,3 ]
McCormack, Matthew [1 ,2 ,3 ]
Sheen, Jen [1 ,2 ,3 ]
机构
[1] Massachusetts Gen Hosp, Dept Mol Biol, Boston, MA 02114 USA
[2] Massachusetts Gen Hosp, Ctr Computat & Integrat Biol, Boston, MA 02114 USA
[3] Harvard Univ, Sch Med, Dept Genet, Boston, MA 02114 USA
来源
PLANT METHODS | 2012年 / 8卷
关键词
Next generation sequencing; EMS; PCR-amplified genomic library; Nitrate signalling; Positional cloning; MAP-BASED CLONING; THALIANA; IDENTIFICATION; NITROGEN; GALAXY; TRANSFORMATION; DNA;
D O I
10.1186/1746-4811-8-12
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Large-scale genetic screens in Arabidopsis are a powerful approach for molecular dissection of complex signaling networks. However, map-based cloning can be time-consuming or even hampered due to low chromosomal recombination. Current strategies using next generation sequencing for molecular identification of mutations require whole genome sequencing and advanced computational devises and skills, which are not readily accessible or affordable to every laboratory. We have developed a streamlined method using parallel massive sequencing for mutant identification in which only targeted regions are sequenced. This targeted parallel sequencing (TPSeq) method is more cost-effective, straightforward enough to be easily done without specialized bioinformatics expertise, and reliable for identifying multiple mutations simultaneously. Here, we demonstrate its use by identifying three novel nitrate-signaling mutants in Arabidopsis.
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页数:12
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