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PolyA-selected mRNA was prepared from unfertilized Xenopus laevis eggs. The library was constructed in the vector pT7T3-Pac as described in Bonaldo, M.F., Lennon, G. and Soares, M.B. 'Normalization and subtraction: two approaches to facilitate gene discovery', Genome Research 6:791-806, 1996. The first strand synthesis used a NotI-dT18 primer; double stranded cDNAs were ligated to EcoRI adapters, digested with NotI, and directionally cloned into the NotI and EcoRI-digested pT7T3-Pac vector. The library contained approximately 7.2 X 10^5 recombinants, with average insert sizes of 1-1.5 kb.
Nucleotide
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