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CHORI-561: H9 Human Embryonic Stem cell line BAC Library |
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The CHORI-561 H9 Human Embryonic Stem cell line BAC Library was constructed from a suspension of H9 embryonic stem cells. The library was generated by Yongli Bai and Maxim Koriabine in Pieter de Jong’s laboratory at BACPAC Resources, Children’s Hospital Oakland Research Institute. Procedures followed the cloning approach developed in our laboratory (Osoegawa et al., 1998). The cells were provided by Dr. James Thomson, University of Wisconsin Stem Cell & Regenerative Medicine Center. The cell suspension was embedding in 0.5% InCert agarose and proteins were removed by a detergent/proteinaseK treatment. The resulting high molecular weight DNA was partially digested using a combination of EcoRI restriction enzyme and EcoRI methylase enzyme then size fractionated by pulsed-field gel electrophoresis. DNA fragments from the appropriate size fraction were cloned into the pSG1473 vector between the two EcoRI sites. The ligation products were transformed into MDS42recA trfA Blue electro-competent cells (Scarab Genomics). The library has been arrayed into 624 (384-well) microtiter dishes.
Segment | 1 | 2 | All | Vector | pSG1473 | pSG1473 | pSG1473 | Restriction Enzyme | EcoRI/EcoRI Methylase | EcoRI/EcoRI Methylase | EcoRI/EcoRI Methylase | DNA Source | cell line | cell line | cell line | Plate Numbers | 1-336 | 337-624 | 1-624 | Plate Count | 336 | 288 | 624 | Empty Wells | N/A | N/A | 2204 (0.92%) | Non-Recombinant Clones | N/A | N/A | N/A | Non-Insert Clones | N/A | N/A | Approx. 3158 (1.33%) | Recombinant Clones | 129024 | 110592 | 234254 | Average Insert Size | N/A | N/A | 172 Kbp | Genomic Coverage | N/A | N/A | N/A |
Data on the CHORI-561 clone insert size distribution is currently not available for this library and will be posted as soon as available.
No clones were found to be non-recombinant after analysis of CHORI-561 using overgo probes specific for the puc19 fragment.
Question related to this library should be sent to Pieter J. de Jong ([email protected], fax: (510) 450-7924).
Posted: October 17, 2007
Revised: April 8, 2008
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