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Human Universal cDNA Library Array I

used for in situ hybridization on colonies. The hybridization signals are detected by autoradiography or phosphorimaging, and the positive pools are easily located by rows and columns printed on the membranes (Figure 6). When the position of a positive pool has been identified, the corresponding Secondary Membrane gridded with the individual colonies can be obtained from Stratagene for a secondary screening. The result of thesecondary screening determines the exact location of the target cDNA clone, which can then be obtained as a bacterial stab culture.

Conclusions

Stratagenes Human Universal cDNA Library Array I represents a collection of over 145,000 primary clones individually organized in microwell plates and stored at -80C. The high quality of these clones obtained after extensive normalization has been demonstrated by an average insert length of 1.7 kb, a representation of the ubiquitous gene b-actin 25- to 100-fold lower than in the direct library, and a high representation of full-length clones (up to 70% for b-actin). Using this material, Stratagene has developed a powerful tool to accelerate gene discovery by nucleic acid screening: A first set of four Master Membranes allows a rapid screening of pools and can be reused for different cloning projects, while a Secondary Membrane is used to identify the final position of the target cDNA clone. Human Universal cDNA Library Array I allows researchers to efficiently identify and isolate human genes by screening a large representation of high-quality cDNA clones in a short period of time.

Methods

HUCL Array I Master Membranes were hybridized using a conventional method. Prehybridizatio
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