Cloning and partial sequencing of a new rat brain specific cDNA

Abstract

In order to find brain specific transcripts in a pUC13 cDNA library prepared from rat brain cytoplasmic poly(A)⁺ RNAs, the following steps were observed: 1. Randomly chosen cDNA clones from the brain library were screened by radiolabell-ed single stranded cDNAs (sscDNAs) prepared from liver, spleen, kidney and intestine mRNAs. 2. The brain clones containing genetic information shared with the peripheral organs were discarded. 3. After hybridization of the remaining clones with radiolabeled sscDNAs prepared from rat brain poly(A)⁺ RNAs, 210 possibly brain specific clones were selected. 4. Plasmids containing cDNA of each of these clones were purified and after estimation of their cDNA sizes, clones containing inserts of at least 500 base pairs (bp) were used as radiolabeled probes against rat brain, liver, spleen, kidney and intestine po-ly(A)⁺ RNAs spotted onto nitrocellulose filters (dot blot analysis). 5. By using this screening procedure, several brain specific clones were obtained. One of them (clone III 25) is presently subjected to sequencing in Ml3 phage. 6. A computer analysis of the partial sequence so far obtained gives no significant homology to the 15.000 genes contained in Genebank and EMBL data bank. It seems therefore that we have isolated a new brain specific cDNA coding for a not yet described peptide.