|
CHORI-256: Galago (Ottolemur garnetti) BAC Library |
 |
The CHORI-256 Galago (Otolemur garnetti, male) BAC Library has been generated by Doriana Misceo and Dr. Baoli Zhu, in Pieter de Jong’s laboratory at BACPAC Resources, Children’s Hospital Oakland Research Institute. The library preparation followed the general cloning approach developed in our laboratory Osoegawa et al., 1998. Kidney tissue was obtained from a male galago provided by the Department of Cell & Developmental Biology, Vanderbilt Medical School. The frozen kidney was ground into a powder, cells embedded in 0.5% InCert agarose then proteins were removed by a detergent/proteinaseK treatment. The resulting agarose-embedded high molecular weight DNA was partially digested with a combination of EcoRI restriction enzyme and EcoRI methylase then size fractionated by pulsed-field gel 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 has been arrayed into 576, 384-well microtiter dishes and has subsequently been gridded onto 12, 22x22cm nylon high-density filters for screening by probe hybridization. Each hybridization membrane represents over 18,000 distinct BAC clones, stamped in duplicate. The BAC library construction was funded by a grant from the NIH (#HG01165-06).
Data for CHORI-256 Galago 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 | 289-576 | 1-576 | Plate Count | 288 | 288 | 576 | Empty Wells | N/A | N/A | N/A | Non-Recombinant Clones | N/A | N/A | N/A | Non-Insert Clones | Approx. 221 (0.2%) | Approx. 221 (0.2%) | Approx. 885 (0.4%) | Recombinant Clones | 110371 | 110371 | 220299 | Average Insert Size | 176 Kbp | 176 Kbp | 176 Kbp | Genomic Coverage | 5X | 5X | 10X |
Click here for legend of the previous table.
No clones were found to be non-recombinant after analysis of CHORI-256 using overgo probes specific for the puc19 fragment.
Data on the CHORI-256 clone insert size distribution is currently not available for this library and will be posted as soon as available.
The library is available for individual clone orders distributed as agar-stab cultures in a fee structure described on our website "Ordering & Pricing Information" .
The BAC library has been gridded onto 22x22cm positively charged nylon filters for hybridization screening purposes. Each filter contains 36,864 colonies which represents 18,432 independent clones spotted in duplicate in a 4x4 clone array. Clones can be identified through screening by purchasing "high-density colony" hybridization filters or by utilizing our "fee for service" "Library Screening Services" . For questions about clones and hybridization membranes, please contact BACPAC Resources ([email protected]).
Please direct questions concerning this library to either Pieter J. de Jong.The library is available to academic users on a cost recover basis, reimbursement of duplication costs (labor & materials). Dishes can only be obtained per complete library or per segment of the library. Individual clones identified after screening can be obtained as stab cultures "Ordering & Pricing Information" .
Shipping will be done by Federal Express and charged to the recipients account number. With respect to the scheduling of shipments of arrayed library copies, we would like to make you aware of our policy for array distribution.
Content Posted: Sept. 30, 2003
Content Revised: Sept. 14, 2007
|
