Construction and characterization of a 10-genome equivalent yeast artificial chromosome library for the laboratory rat, Rattus norvegicus.
Increasing attention has been focused in recent years on the rat as a model organism for genetic studies, in particular for the investigation of complex traits, but progress has been limited by the lack of availability of large-insert genomic libraries. Here, we report the construction and characterization of an arrayed yeast artificial chromosome (YAC) library for the rat genome containing approximately 40,000 clones in the AB1380 host using the pCGS966 vector. An average size of 736 kb was estimated from 166 randomly chosen clones; thus the library provides 10-fold coverage of the genome, with a 99.99% probability of containing a unique sequence. Eight of 39 YACs analyzed by fluorescence in situ hybridization were found to be chimeric, indicating a proportion of about 20-30% of chimeric clones. The library was spotted on high-density filters to allow the identification of YAC clones by hybridization and was pooled using a 3-dimensional scheme for screening by PCR. Among 48 probes used to screen the library, an average of 9.3 positive clones were found, consistent with the calculated 10-fold genomic coverage of the library. This YAC library represents the first large-insert genomic library for the rat. It will be made available to the research community at large as an important new resource for complex genome analysis in this species.
Cai
L
L
Cardiovascular Division, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA.
Schalkwyk
L C
LC
Schoeberlein-Stehli
A
A
Zee
R Y
RY
Smith
A
A
Haaf
T
T
Georges
M
M
Lehrach
H
H
Lindpaintner
K
K
D000818:Animals; D018244:Chromosomes, Artificial, Yeast; D004247:DNA; D015698:Genomic Library; D017404:In Situ Hybridization, Fluorescence; D051381:Rats
D004247:DNA