INRA/UMR Biologie du Developpement et Reproduction
Manufacture protocol
The 10K array contains 7.800 EST from term placenta (Everts et al., 2005), incremented by 2.400 EST from the BE03 cDNA library. The BE03 library was established on extra-embryonic tissues of bovine embryos from D14 to D24 of gestation and fetuses (D36 and D64 and has been indexed in Unigene as Lib. 17188 and Lib.15992. In total, 10.214 unique cDNA were spotted onto Nylon N+ membranes (Amersham Biosciences) with a 3x3 pattern (QBot, Proteigene) at the CRB GADIE platform (INRA, Jouy en Josas, France).
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nylon N+ membranes
Description
(22cmx22cm, Amersham Biosciences) - 3x3 pattern - The 10K custom cDNA membrane developed in our laboratory was used for the experiment. Briefly, the array contained 7.800 EST from term placenta (Everts et al., 2005) and we incremented this set by 2.400 new EST from a cDNA library established on young foetuses and early extra-embryonic tissues. This second library (called BE03) was constructed from mRNA isolated from extra-embryonic tissues of bovine embryos from D14 to D24 of gestation and foetuses (D36 and D64). The BE03 library was normalized and subtracted by the 7.8K set using the protocol from Bento Soares. About 10.000 bacterial clones were sequenced. The sequences were repeat-masked using RepeatMasker (05/05/99) and the clustering was performed by CAP3. The CAP3 assembly resulted into 2.400 new EST. In total, 10.214 unique cDNA were spotted onto Nylon N+ membranes (Amersham Biosciences) with a 3x3 pattern (QBot, Proteigene) at the CRB GADIE platform (INRA, Jouy en Josas, France). The “BE03” library is indexed as Lib. 17188 and Lib. 15992 (http://www.ncbi.nlm.nih.gov).
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