Chapter title |
Genomic Libraries: II. Subcloning, Sequencing, and Assembling Large-Insert Genomic DNA Clones.
|
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Chapter number | 4 |
Book title |
Molecular Methods for Evolutionary Genetics
|
Published in |
Methods in molecular biology, July 2011
|
DOI | 10.1007/978-1-61779-228-1_4 |
Pubmed ID | |
Book ISBNs |
978-1-61779-227-4, 978-1-61779-228-1
|
Authors |
Quail MA, Matthews L, Sims S, Lloyd C, Beasley H, Baxter SW, Quail, Mike A., Matthews, Lucy, Sims, Sarah, Lloyd, Christine, Beasley, Helen, Baxter, Simon W., Mike A. Quail, Lucy Matthews, Sarah Sims, Christine Lloyd, Helen Beasley, Simon W. Baxter |
Abstract |
Sequencing large insert clones to completion is useful for characterizing specific genomic regions, identifying haplotypes, and closing gaps in whole genome sequencing projects. Despite being a standard technique in molecular laboratories, DNA sequencing using the Sanger method can be highly problematic when complex secondary structures or sequence repeats are encountered in genomic clones. Here, we describe methods to isolate DNA from a large insert clone (fosmid or BAC), subclone the sample, and sequence the region to the highest industry standard. Troubleshooting solutions for sequencing difficult templates are discussed. |
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