SHEPHERD%ESVAX@dupont.com (04/03/91)
The chromosome 22 bulletin board has been in existence for some time now
with remarkably little effort to use it. I would like to try and break
that trend by posting a brief paragraph concerning some on-going research
and a request for some specific help. I urge each of you to do the same so
that we will become more familiar with those persons actually monitoring
the bulletin board. In this way, the bulletin board can really become a
tool for facilitating research on chromosome 22.
I am an investigator in the newly formed DuPont Merck Pharmaceutical
Company in Wilmington Delaware. As a collaborative effort with several
colleagues and persons at other institutions, I have become involved
in the effort to map human chromosome 22. Specifically, we would like
to construct a chromosome 22 genomic library in
the bacteriophage P1 vector cloning system (a novel cloning system that
allows the cloning of 70-100 kb inserts, developed at duPont by Nat
Sternberg). The input DNA for this chromosome specific library will be
flow-sorted material. In addition, we are interested in arraying a 3-5X
total human/P1 genomic library as individual clones. Chromosome-22 clones
can, of course, also be derived from this library. The P1 clones will be
used in conjunction with chromosome-22 YAC clones (prepared by others) to
try and get a complete coverage of chromosome 22.
Specific items that I would like to get a response to are:
1. We currently use Cot1 human DNA for cold competitor DNA when using a
specific between Alu PCR fragment as a hybridization probe. This works
fine. However, to screen the total human library for chromosome 22
specific clones, we would like to use as a hybridization probe the total
mixture of between Alu PCR products that is derived from a CHO/22
hybrid(s). In trying this, we find too many colonies hybridizing,
suggesting that for some reason the competition of repetitive
sequences is not working. If you have worked out the conditions
for doing this particular type of probing, I would very much
appreciate the information.
2. If you have experience in the preparation and cloning from flow-sorted
chromosome 22 material, I would appreciate getting some detailed comments
from you.
3. If you have knowledge of someone that has developed an automated method
of picking colonies from a random array on a plate into an ordered array in
microtiter wells, I would appreciate a response.
4. If you are aware of a good -70C storage-shelving system for
bacterial glycerol stocks in microtiter well dishes (preferably
for an upright freezer), let me know.
THANKS a lot in advance. May I also suggest that you respond to the
entire chromosome-22 bulletin board? (See below)
Location Posting Address
-------------------------------------------------------
Americas / Internet format: chromosome-22@genbank.bio.net
Americas / BITNET format: chrom-22@genbank.bio.net
Ireland / EARN format: chrom-22@irlearn.ucd.ie
U.K. / JANET format: chrom-22@uk.ac.daresbury
Sweden / Internet format: chromosome-22@bmc.uu.se
Nancy Shepherd, PhD
The DuPont Merck Pharmaceutical Company
E328/148B
Wilmington, Delaware 19880-0328
phone 302-695-1519
fax 302-695-7556
"Shepherd%ESVAX@dupont.com"