| CHORI-258
Owl Monkey BAC Library |
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Provisonal web page, posted June 9, 2003:
This library is not yet available and is scheduled to become
accessible for filter and clone orders per June 30, 2003 (orders
accepted in advance). Library copies will be prepared once
per year upon request – to maintain clone viability
in the master array. The first replication will take place
during September 2003 for orders received prior to September
1, 2003.
Background information:
Background information: The CHORI-258 Owl Monkey BAC library
has been constructed by Baoli Zhu in Pieter de Jong’s
laboratory at BACPAC Resources, Children’s Hospital
Oakland Research Institute. The preparation of the library
followed the cloning approach developed in our laboratory
(Osoegawa
et al., 1998). DNA was isolated from kidney cells obtained
from a male owl monkey (Aotus nancymai) obtained from The
Squirrel Monkey
Breeding and Research Resource (SMBRR, Department of Comparative
Medicine, University of South Alabama, Mobile, AL36688). The
local SMBRR identification number for this animal is 85286.
A skin biopsy and a blood sample have been sent to the Coriell
Cell Repositories to establish cell lines as part of the Integrated
Primate Biomaterials and Information Resource (IPBIR)
initiative. High-molecular-weight DNA was isolated from a
kidney cell suspension prepared in a Dounce Homogenizer and
was embedded in agarose to stabilize the DNA. The 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 528 384-well microtiter dishes and will
subsequently be gridded onto 11 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 (#HG025323-02).
Provisional Data for Owl Monkey BAC library:
| Segment |
1 |
2 |
Total |
| Cloning Vector |
pTARBAC2.1 |
pTARBAC2.1 |
|
| DNA Source |
Kidney |
Kidney |
|
| Restriction Enzyme |
EcoRI |
EcoRI |
|
| Plate Range |
1-288 |
289-528 |
1-528 |
| Total 384-Well Dishes |
288 |
240 |
528 |
| Empty Wells (%) |
N/A |
N/A |
N/A |
| Non-insert Clones (%) |
N/A |
N/A |
N/A |
| Vector Clones (%) |
N/A |
N/A |
N/A |
| Recombinant Clones |
N/A |
N/A |
N/A |
| Insert size (Average) |
~165 Kbp |
~165 Kbp |
~165Kbp |
| Standard Deviation |
N/A |
N/A |
N/A |
| Library Redundancy (3.3 Gbp genome) |
~4.6 fold |
~5.5 fold |
~10.1 fold |
click here for a legend of
the previous tables.
The CHORI-258 clone average insert size has been determined
from a sample of 172 colonies prior to the arraying of the
BAC library. Pulsed Field Gel Electrophoresis of Not1 digested
BACs with appropriate size standard was used for size determination.
The library will be available as individual clones, and high-density
hybridization filters once the arraying, library duplication
and initial characterization is complete.
Individual clones
identified after screening can be obtained as stab cultures
at a cost of $80 per clone in addition to a $35 set-up charge
per shipment. 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.
Please visit the ordering information
page if you want to place an order.
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). For hybridization
membranes, please contact BACPAC Resources (BACPACorders@chori.org).
Date initially posted: June 9, 2003. Revision dates: June
9, 2003; June 19th, 2003
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