Stability of messenger RNA in postmortem human brains and construction of human brain cDNA libraries.
We studied stabilities of poly(A)(+)-RNA in postmortem mouse and human brains for up to 12 hours. The yields of total RNA were not changed significantly during postmortem periods either in mouse brains or human brains. Cell-specific cDNA probes were used to evaluate postmortem stability of poly(A)(+)-RNA in each cell type in the central nervous system. We used neuron-specific enolase (NSE), S-100 beta (S-100), and myelin-associated glycoprotein (MAG) for molecular markers of neuron, astrocyte, and oligodendrocyte, respectively. There was no detectable degradation of mRNAs coding for NSE, S-100, and MAG during the postmortem periods on Northern blot hybridization analyses. These results indicate that intact mRNAs expressed in neuron, astrocyte, or oligodendrocyte can be isolated from postmortem brains for up to 12 hours after death. Using poly(A)(+)-RNA thus isolated from two postmortem human brains, we constructed directional cDNA libraries and demonstrated the presence of full-length cDNAs for NSE, S-100, and MAG on Southern blot hybridization analysis. The present data should encourage studies on altered gene expressions in human brain in various neurologic diseases.
Department of Neurology, Niigata University, Japan.
H
H
Kobayashi
K
K
Sakimura
R
R
Kuwano
S
S
Sato
F
F
Ikuta
Y
Y
Takahashi
T
T
Miyatake
S
S
Tsuji
D000368:Aged; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D001923:Brain Chemistry; D015342:DNA Probes; D005260:Female; D015723:Gene Library; D006801:Humans; D051379:Mice; D008875:Middle Aged; D008969:Molecular Sequence Data; D009185:Myelin Proteins / Q000235:genetics; D018992:Myelin-Associated Glycoprotein; D010751:Phosphopyruvate Hydratase / Q000235:genetics; D011061:Poly A / Q000737:chemistry*; D011180:Postmortem Changes; D012333:RNA, Messenger / Q000737:chemistry*; D009418:S100 Proteins / Q000235:genetics
D015342:DNA Probes; D009185:Myelin Proteins; D018992:Myelin-Associated Glycoprotein; D012333:RNA, Messenger; D009418:S100 Proteins; D011061:Poly A; D010751:Phosphopyruvate Hydratase