Repeat subtraction-mediated sequence capture from a complex genome
Article first published online: 4 MAR 2010
© 2010 The Authors. Journal compilation © 2010 Blackwell Publishing Ltd
The Plant Journal
Volume 62, Issue 5, pages 898–909, June 2010
How to Cite
Fu, Y., Springer, N. M., Gerhardt, D. J., Ying, K., Yeh, C.-T., Wu, W., Swanson-Wagner, R., D’Ascenzo, M., Millard, T., Freeberg, L., Aoyama, N., Kitzman, J., Burgess, D., Richmond, T., Albert, T. J., Barbazuk, W. B., Jeddeloh, J. A. and Schnable, P. S. (2010), Repeat subtraction-mediated sequence capture from a complex genome. The Plant Journal, 62: 898–909. doi: 10.1111/j.1365-313X.2010.04196.x
- Issue published online: 25 MAY 2010
- Article first published online: 4 MAR 2010
- Received 30 October 2009; revised 3 February 2010; accepted 9 February 2010; published online 12 April 2010.
- NimbleGen sequence capture;
- molecular marker;
- reduced representation sequencing;
- allele mining
Sequence capture technologies, pioneered in mammalian genomes, enable the resequencing of targeted genomic regions. Most capture protocols require blocking DNA, the production of which in large quantities can prove challenging. A blocker-free, two-stage capture protocol was developed using NimbleGen arrays. The first capture depletes the library of repetitive sequences, while the second enriches for target loci. This strategy was used to resequence non-repetitive portions of an approximately 2.2 Mb chromosomal interval and a set of 43 genes dispersed in the 2.3 Gb maize genome. This approach achieved approximately 1800–3000-fold enrichment and 80–98% coverage of targeted bases. More than 2500 SNPs were identified in target genes. Low rates of false-positive SNP predictions were obtained, even in the presence of captured paralogous sequences. Importantly, it was possible to recover novel sequences from non-reference alleles. The ability to design novel repeat-subtraction and target capture arrays makes this technology accessible in any species.