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CHORI-273: Black Lemur (F) (Eulemur macaco macaco) BAC Library |
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The CHORI-273 Black Lemur (Eulemur macaco macaco) BAC library has been constructed by Boudewijn ten Hallers and Baoli Zhu in Pieter de Jong’s laboratory at BACPAC Resources, Children’s Hospital Oakland Research Institute. The library preparation followed the cloning approach developed in our laboratory (Osoegawa et al., 1998). Duke University Primate Center provided the kidney tissue from a female black lemur. (Additional data about the black lemur individual: Emm, Pandora II; DUPC ID# 6358 female; Date of Birth: 3/26/1990; Date of Death: 5/14/1997; Cause of death: euthanasia due to fibrosarcoma; Tissue obtained upon necropsy, and frozen at -80C in approximately 1 hour.)
High-molecular-weight DNA was isolated from frozen kidney tissue and was embedded in agarose to stabilize the DNA. Agarose embedded DNA was partially digested with a combination of EcoRI restriction enzyme and EcoRI methylase and size fractionated by pulsed-field electrophoresis. DNA fragments from the appropriate size fraction were cloned into the pTARBAC2.1 vector between the two EcoRI sites. The ligation products were transformed into DH10B (T1 resistant) electro-competent cells (Invitrogen). The library will be arrayed into 576 (384-well) microtiter dishes and will subsequently be gridded onto twelve 22x22cm nylon high-density filters for screening by probe hybridization. Each hybridization membrane represents over 18,000 distinct BAC clones, stamped in duplicate.
BAC library construction was funded as part of the NIH-funded BAC Resource Network by a grant from NIH.
Provisional Data for CHORI-273 Black Lemur (Eulemur macaco macaco) BAC library:
Segment | 1 | 2 | All | Vector | pTARBAC2.1 | pTARBAC2.1 | pTARBAC2.1 | Restriction Enzyme | EcoRI/EcoRI Methylase | EcoRI/EcoRI Methylase | EcoRI/EcoRI Methylase | DNA Source | kidney | kidney | kidney | Plate Numbers | 1-288 | 289-528 | 1-528 | Plate Count | 288 | 240 | 528 | Empty Wells | 3423 (3.1%) | 2351 (2.55%) | 5774 (2.85%) | Non-Recombinant Clones | 0 (0%) | 0 (0%) | 0 (0%) | Non-Insert Clones | Approx. 1629 (4/264, 1.52%) | Approx. 1590 (4/226, 1.77%) | Approx. 3211 (8/490, 1.63%) | Recombinant Clones | 105540 | 88219 | 193767 | Average Insert Size | 195 Kbp | 185 Kbp | 190 Kbp | Genomic Coverage | 5.5X | 4.5X | 10X |
Data on the CHORI-273 clone average insert size has been determined by Pulsed Field Gel Electrophoresis of NotI digested BACs. Clone size distribution graph will be posted on this page as soon as available.
Further in depth characterization of the library is on going in our lab and data will be updated on our web page periodically.
Please direct questions concerning this library to either Pieter J. de Jong.
Ordering & Pricing information
The library is available in several formats. Individual clones, and high-density hybridization filter are obtainable. For ordering and shipping details, please view the ordering and pricing information ordering and pricing information page.
Academic and commercial users interested in a copy of the BAC library should contact Pieter J. de Jong ([email protected], fax: (510) 450-7924).
Reference
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