LLNL HUMAN SINGLE CHROMOSOME COSMID LIBRARY :
Chromosome 2
LL02NP04
"AI"
About the library
This Chromosome 2 PAC library was constructed at LLNL. It has an average
insert size of 85 Kb and contains 5,760 clones providing a 1.9-fold coverage
of the chromosome. The vector used is pCYPAC2N. The library consists of
15 plates labelled 1-15.
Production of high-density gridded filters
The library has been gridded in a 4x4 array on one 22
x 7.4 cm Hybond N nylon membrane (Amersham). Each clone has been
spotted twice to give 11,520 clones (5,760 x 2) on the membrane
which is labelled "AI".
Clones spotted on the filters were grown overnight at 37°C on LB
agar plates containing Kanamycin (20µg/ml). The filters were subsequently
processed by (a) SDS treatment, (b) denaturation and (c) neutralisation,
then dried and crosslinked using a Spectronics Corporation XL-1500 UV
Crosslinker. The filters are ready for pre-hybridisation in appropriate
buffers.
User Information:
For successful removal of probes, and to enable re-probing, membranes
must NEVER be allowed to dry during, or after, hybridisation and washing.
Membranes can be stripped to facilitate re-probing by washing sequentially
at RT in 0.2M NaOH for 30 min. and 0.1 x SSC / 0.1% SDS / 0.2M Tris-HCl pH 7.5 for 15 min. A gentler method, which extends the number of probings
per filter, is to wash the probed membrane after use in a large volume
of pre-hybridisation buffer overnight at 65°C, prior to using
another probe.
Subsequent publication:
Please acknowledge both the originator of the library, LLNL as
described in the "materials transfer agreement" that
you have signed, and Geneservice for providing
the library, in any publication arising out of using this resource.
An example of how a clone (E.g. CLONE NAME = 9-B9) should be described
in a publication is: AI9b9 from the LL02NP04 library.
5th Sept 2000 Version 1.2
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