Enhanced access to rare brain cDNAs by prescreening libraries: 207 new mouse brain ESTs.
To use single-pass cDNA sequencing to characterize low-frequency cDNA clones from a region of the brain that includes the primary site of neurodegeneration in human Parkinson disease, we have developed a prescreening procedure using single brain region first-strand cDNA probes. Selection of cDNA clones giving low hybridization signals allowed the elimination of clones resulting from abundant messages and enrichment for clones corresponding to low-copy messages. Comparative sequencing of standard and prescreened cDNA libraries (191 and 124 clones, respectively) showed that this procedure raised the frequency of novel sequences encountered from 54 to 81%. The increased proportion of novel ESTs justifies the labor of prescreening. Automation of this procedure will accelerate the molecular description of genes expressed in any brain region, or any tissue, and represents a way to maximize access to cDNA sequences for human and mouse genome characterization. In total, the comparative sequencing experiments generated 207 new mouse and 11 new rat brain ESTs.
Davies
R W
RW
Institute of Biomedical and Life Sciences, Robertson Laboratory of Biotechnology, University of Glasgow, Scotland, United Kingdom.
Roberts
A B
AB
Morris
A J
AJ
Griffith
G W
GW
Jerecìć
J
J
Ghandi
S
S
Kaiser
K
K
Savioz
A
A
D000818:Animals; D001921:Brain / Q000378:metabolism*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D015870:Gene Expression; D015723:Gene Library; D005821:Genetic Techniques; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D051381:Rats; D017423:Sequence Analysis, RNA
D018076:DNA, Complementary