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CHORI-257: Grey Mouse Lemur (F) (Microcebus murinus) BAC Library |
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The CHORI-257 mouse lemur BAC library has been constructed
by Michael Nefedov 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). Kidney was obtained from an adult female
mouse lemur (Microcebus murinus) through John Allman, Division
of Biology 216-76, Caltech, Pasadena, CA 91125. The local
identification number for this animal is #8. 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
the cells were 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 has been arrayed into 576 “384-well”
microtiter dishes and also gridded onto twelve 22x22cm
nylon high-density filters for screening by probe hybridization.
Each hybridization membrane represents about 18,000 distinct
mouse lemur BAC clones, stamped in duplicate. Library characterization
was performed by Qing Cao, Teresa Ren and Kazutoyo Osoegawa.
This work was funded by NIH grant HG025323-01.
| Segment |
1 |
2 |
Total |
| Cloning Vector |
pTARBAC2.1 |
pTARBAC2.1 |
|
| DNA Source |
Kidney Cell |
Kidney Cell |
|
| Restriction Enzyme |
EcoRI |
EcoRI |
|
| Plate Range |
1-288 |
289-576 |
1-576 |
| Total 384-Well Dishes |
288 |
288 |
576 |
| Empty Wells (%) |
1,000 (0.9%) |
2,400 (2.2%) |
3,500 (1.5%) |
| Non-insert Clones (%) |
4/287 (1.4%) |
7/274 (2.6%) |
11/561 (2.0%) |
| Vector Clones (%) |
0/110K (0) |
0/110K (0) |
0/220K (0) |
| Recombinant Clones |
~108K |
~105K |
~213K (>96% of wells) |
| Insert size (Average) |
~210Kbp |
~220Kbp |
~214Kbp |
| Standard Deviation |
57Kbp |
55Kbp |
56Kbp |
| Library Redundancy |
~7.6 fold |
~7.7 fold |
~15 fold |
Click here for a legend of
the previous tables.
Data on the CHORI-257 clone average insert size has been determined
by Pulsed Field Gel Electrophoresis. Clone size distribution
has been plotted graphically.
Dishes can be obtained per
complete library ($11,520.00) or per segment ($5,760.00).
Hybridization filters can be ordered per complete set of 12
membranes (price per set is 12 times $200.00) or per segment
(6 times $200.00). Shipping will be done by Federal Express
and charged to recipient's 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 ([email protected],
fax: (510) 450-7924). For hybridization
membranes, please contact BACPAC Resources ([email protected]).
Please contact Pieter
J. de Jong for questions related to this library.
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