Several labs have been using this l ZAP Hydra vulgaris cDNA library made by Stratagene (and generously paid for by Mike Sarras, University of Kansas Medical Center). Others may want to use this library. Here is all of the information we have on the library. Requests for aliquots of the library should be directed to Hans Bode at UCI.
The library is a Stratagene custom cDNA library made from polyA+ RNA from whole adult polyps of Hydra vulgaris (UCI strain) by a combination of oligo-dT and random oligo priming. The cDNA was cloned into the Stratagene vector l ZAPII. 1.1 x 107 primary pfu's were obtained. 1 x 106 of these pfu's were amplified once. The remainder of the primary phage were aliquoted and are stored frozen in the Bode lab. Less than 5% of the primary phage were nonrecombinants. The titer of the amplifed library is 4.0 x 1010 pfu/ml. The amplified library has been aliquoted and is stored frozen in the Bode lab. Adaptors at the ends of inserts are as follows (EcoRI sites are in lower case): gaattcGGCACGAG / cDNA insert / CTCGTGCCgaattc
Some inserts in this library have been found (by digestion and confirmed by DNA sequencing) to have a damaged EcoRI site at one end. In such cases the insert cannot be released by EcoRI cleavage. The insert can, however, be released by a double digest with NotI and XhoI, enzymes whose sites flank the EcoRI site and which are unlikely to cut in Hydra DNA. Inserts at least as large as 3 kb are present in the library. In vivo excision from l ZAPII releases the insert into the phagemid vector pBluescript SK-. Plating of the library and in vivo excision are done in E. coli XL1-Blue.
Clones from this library have been isolated by oligonucleotide screening, antibody screening, and screening with fragments from other clones. Ruth Weinziger (Munich) has found that there are some "hybrid" clones in this library in which cDNA fragments from two unrelated genes have been ligated together and inserted into a single phage clone. Michael Kroiher (Koeln) has found an Artemia mitochondrial DNA clone in the library, so one should be careful about assuming that everything gotten from the library is from Hydra.
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