RPCI-104 Schizosaccharomyces pombe BAC Library
The RPCI-104 BAC Library has been constructed in our laboratory. Genomic DNA was purchased from BIORAD (Cat# 170-3633) and partially digested with a combination of EcoRI and EcoRI Methylase. Size selected DNA was cloned into the pBACe3.6 vector between the EcoRI sites. The ligation products were transformed into DH10B electro-competent cells (BRL LIFE TECHNOLOGIES). The library has been arrayed into 384-well microtiter dishes and also gridded onto 11 x 7cm nylon high-density filters for screening by probe hybridization. Each hybridization membrane represents over 3,000 distinct Schizosaccharomyces pombe BAC clones, stamped in duplicate. Library construction was supported by sub-contracts from a grant awarded to Dr. Kazuei Mita in The National Institute of Radiological Sciences.
The RPCI-104 Schizosaccharomyces pombe BAC Library
|
Segment
|
DNA
|
Restriction Enzyme
|
Plate Numbers
|
Total Plates
|
Empty Wells
|
Empty Wells (%)
|
|
|
1
|
pBACe3.6
|
S.pombe 972H minus (ATCC 24843)
|
EcoRI
|
1-8
|
8
|
16
|
0.5
|
|
Segment
|
Non-insert Clones (%)
|
Non-insert clones
|
Recombinant Clones
|
Insert size (Average)
|
Redundancy (Genome size:
15 Mb) |
|
|
1
|
1.2
|
Approx. 37
|
1
|
3018
|
166Kbp
|
33
|
click here for a legend of the previous tables.
This library should represent approximately 33-fold total genomic representation.
Data on the RPCI-104 clone
average insert size has been determined by Pulsed Field Gel Electrophoresis.
Clone size distribution has been plotted graphically.