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Jörg Großhans cDNA libraries from ovarian RNA for the interaction trap Ovaries of Drosophila melanogaster (OregonR) were prepared according to a mass procedure described in Drosophila, A laboratory manual, CSH Press 1989. RNA was isolated by extraction with guanidinium thiocyanate, followed by CsCl ultrazentrifugation and fractionation with OligodT cellulose. Unidirectional cDNA was synthesized with a ZAP cDNA synthesis kit from Stratagene and fractionated with Sepharose CL-4B (1 ml column). Two fractions (OvoI and OvoII) were collected. The cDNAs with EcoRI and XhoI sites at their ends were ligated into linearized vector pJG4-5. E. coli SURE (Stratagene) was transformed with the ligation mix and each was plated onto ten 24x24 cm LB plates with ampicillin. The colonies were directly collected from the plates and used for plasmid DNA isolation by CsCl density gradient centrifugation. Two preparations were made with the OvoI fraction of cDNAs (OvoI and OvoIb). Table 1: Summary
About 50 µg DNA (more or less) of the original DNA preparation are contained in each of the tubes in the form of an ethanol precipitate. Please contact me, if you need more material or have any trouble with it. Reference: |