This Drosophila melanogaster cosmid library was constructed by Inga Siden-Kiamos at the Institute of Molecular Biology and Biotechnology, Heraklion, Greece and supplied by David Glover from the University of Dundee (Siden-Kiamos. I. et al [1990] Nucl. Acids Res. 18: 6261-6270). It has an estimated average insert size of 35kb and contains approximately 18,000 clones providing a 4-fold coverage of the genome. The vector used is Lorist 6 (Gibson. TJ. et al [1987] Gene 53: 283-286). The library consists of 48 plates labelled 1- 50 (plates 45 and 46 are missing).

Production of high-density gridded filters

The library has been gridded in a 4x4 array on one large Hybond N nylon membrane (Amersham). Each clone has been spotted twice.

* plates 45 and 46 are missing

Clones spotted on the filters were grown overnight at 37°C on LB agar plates containing Kanamycin (25 µg/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. The use of two different Hybond membrane types will not affect subsequent hybridisation.

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 and Geneservice Ltd. for providing the library, in any publication arising out of using this resource. An example of how a clone (E.g. CLONE NAME = 32-B9) should be described in a publication is: 32b9 from the cDROS library. The 96-well co-ordinates (CLONE NAME) should be used in all publications.

Version 1.1 17^th August 2000