Comparison of methods for genomic localization of gene trap sequences.
BACKGROUND
Gene knockouts in a model organism such as mouse provide a valuable resource for the study of basic biology and human disease. Determining which gene has been inactivated by an untargeted gene trapping event poses a challenging annotation problem because gene trap sequence tags, which represent sequence near the vector insertion site of a trapped gene, are typically short and often contain unresolved residues. To understand better the localization of these sequences on the mouse genome, we compared stand-alone versions of the alignment programs BLAT, SSAHA, and MegaBLAST. A set of 3,369 sequence tags was aligned to build 34 of the mouse genome using default parameters for each algorithm. Known genome coordinates for the cognate set of full-length genes (1,659 sequences) were used to evaluate localization results.


RESULTS
In general, all three programs performed well in terms of localizing sequences to a general region of the genome, with only relatively subtle errors identified for a small proportion of the sequence tags. However, large differences in performance were noted with regard to correctly identifying exon boundaries. BLAT correctly identified the vast majority of exon boundaries, while SSAHA and MegaBLAST missed the majority of exon boundaries. SSAHA consistently reported the fewest false positives and is the fastest algorithm. MegaBLAST was comparable to BLAT in speed, but was the most susceptible to localizing sequence tags incorrectly to pseudogenes.


CONCLUSIONS
The differences in performance for sequence tags and full-length reference sequences were surprisingly small. Characteristic variations in localization results for each program were noted that affect the localization of sequence at exon boundaries, in particular.
Harper
Courtney A
CA
Department of Biopharmaceutical Sciences, University of California San Francisco, 1700 4th Street, San Francisco, CA 94143-2250, USA. charper@gmail.com
Huang
Conrad C
CC
Stryke
Doug
D
Kawamoto
Michiko
M
Ferrin
Thomas E
TE
Babbitt
Patricia C
PC
D000465:Algorithms; D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics; D056504:Chromatin Assembly Factor-1; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics; D002874:Chromosome Mapping / Q000379:methods; D019295:Computational Biology / Q000379:methods*; D004268:DNA-Binding Proteins / Q000235:genetics; D005091:Exons / Q000235:genetics; D016678:Genome / Q000235:genetics*; D051379:Mice; D009687:Nuclear Proteins / Q000235:genetics; D009711:Nucleotides / Q000235:genetics; D011544:Pseudogenes / Q000235:genetics; D015203:Reproducibility of Results; D016415:Sequence Alignment / Q000379:methods*; D013997:Time Factors
D018797:Cell Cycle Proteins; D056504:Chromatin Assembly Factor-1; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; C501385:Mad1l1 protein, mouse; D009687:Nuclear Proteins; D009711:Nucleotides