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 Home > Resources > Libraries
  CHORI-230 Rat (BN/SsNHsd/MCW) BAC library

  
  Background information

The CHORI-230 BAC Library has been constructed at the Children’s Hospital Oakland Research Institute, BACPAC Resources by Drs Kazutoyo Osoegawa and Baoli Zhu using the cloning techniques developed in our laboratory (Osoegawa et al., 1998). BAC library construction was funded by a grant from NIH (HG01165-06). Rat brain genomic DNA was isolated and partially digested with a combination of EcoRI and EcoRI Methylase. Size selected DNA was cloned into the pTARBAC2.1 vector between the EcoRI sites. For segments 2 of the library, agarose embedded high molecular weight DNA was partially digested by MboI followed by double size fractionation on CHEF gel. Size fractionated DNA was cloned into the pTARBAC1.3 vector between the BamHI sites. The ligation products were transformed into DH10B (T1 resistant) electrocompetent cells (BRL Life Technologies). The library has been arrayed into 384-well microtiter dishes and also gridded onto 22x22cm nylon high-density filters for screening by probe hybridization. Each hybridization membrane represents over 18,000 distinct rat BAC clones, stamped in duplicate.

Rat Source: Howard Jacob's laboratory (Medical College of Wisconsin). DNA was isolated from a 3-month old female BN rat by Kazutoyo Osoegawa during a visit to Dr. Howard Jacobs' lab in September 2000. The complete strain nomenclature is BN/SsNHsd/MCW. BN stands for Brown Norway. SsNHsd is a colony that was obtained from a nucleus colony of Harlan Sprague Dawley (Hsd), at the NIH. The rat BN/SsNHsd was brought to the Medical College of Wisconsin (MCW) 17 generations ago. The rat used for construction of the library is the 14th generation. Due to the incomplete inbreeding of the original Hsd rats, Jacob's lab took care of 17 additional generations of inbreeding towards more complete homozygosity. A general idea of the strain, microsatellite characterization and further links of interest can be obtained from the following URL: http://rgd.mcw.edu

The CHORI-230 BAC Library:

Segment
DNA
Restriction enzyme
Plate Numbers
Total Plates
Empty Wells
Empty Wells (%)
1
pTARBAC2.1
Brain
EcoRI
1-240
240
1,742
1.9
2
pTARBAC1.3
Liver
MboI
241-528
288
1,677
1.5
Total Library
 
 
 
1-528
528
3,419
1.7

 

Segment
Non-insert Clones (%)
Non-insert clones
Recombinant Clones
Average insert size
Redundancy (Genome size: 3,000 Mb)
1
1.1
approx. 995
0
Approx. 89,423
214 Kbp
6.4
2
6.8
7,406
2
Approx. 101,507
162 Kbp
5.5
Total Library
 4.2
8,401
2
Approx. 190,930
186 Kbp 
11.9

 

Data on the CHORI-230 clone average insert size has been determined by Pulsed Field Gel Electrophoresis. Clone size distribution has been plotted graphically. While analyzing clones using pulse-field electrophoresis to determine the average insert size, non-insert clones containing a small deleted vector fragment consistent with sucrose resistance were observed. Further in depth characterization of the library is on going in our lab and data will be updated on our web page periodically.

The library is available in several formats. Individual clones, and high-density hybridization filters are obtainable. For ordering and shipping details, please view the ordering and pricing information  page.

Academic and commercial users interested in a copy of the BAC library should contact Pieter J. de Jong ( pdejong@chori.org ), fax: (510) 450-7924).


Please direct questions concerning this library to either Pieter J. de Jong or Kazutoyo Osoegawa .

 

 

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