Optimisation of methods for selecting candidate genes from cDNA array screens: application to rat brain punches and pineal.
DNA arrays are potentially powerful experimental tools within neuroscience but application of this technology to in vivo paradigms may, in practice, be limited by the sensitivity of transcript detection and inter-screen variation. Here we describe the use of brain punch micro-sampling, used in combination with commercially available cDNA arrays, for profiling brain gene expression in a mutant strain of rat (GAERS model of absence epilepsy). Furthermore, we describe a multi-step optimisation of analysis methods which provides for improved sensitivity and absence of bias in the selection of candidate genes which may be differentially expressed in the mutant. Our method has been validated through application to a second paradigm, rhythmic gene expression in the rat pineal gland. Our experimental design, and analysis method should therefore be generally applicable to subtle discriminations of transcript abundance within discrete brain areas.
Holter
J L
JL
School of Biosciences, Cardiff University, PO Box 911, CF10 3US, Cardiff, UK.
Humphries
A
A
Crunelli
V
V
Carter
D A
DA
D000818:Animals; D001921:Brain / Q000378:metabolism* / Q000503:physiopathology; D001923:Brain Chemistry / Q000235:genetics*; D002940:Circadian Rhythm / Q000235:genetics; D004827:Epilepsy / Q000235:genetics; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation / Q000235:genetics*; D008297:Male; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010870:Pineal Gland / Q000378:metabolism* / Q000503:physiopathology; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011922:Rats, Mutant Strains; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results
D012333:RNA, Messenger