Isolation of high spectral quality RNA using run-on gene transcription; application to gene expression profiling of human brain.
1. Gene expression studies of human brain tissues in both health and disease require the use of intact, high spectral quality messenger RNA (mRNA). Short post-mortem tissues, when adequately controlled, provide a valuable primary source of brain mRNA. 2. Typical mRNA isolation methodologies rely on total RNA extraction from whole cells or tissues. Here we report that as an alternative approach, nuclei can be isolated from the same tissues and programmed for run-on gene transcription to generate enriched mRNA fractions. 3. This novel technique represents a significant improvement over previous methods and provides high quality, high yield mRNA samples suitable for downstream applications that include cloning, sequencing and gene expression studies using DNA array technologies.
Cui
Jian-Guo
JG
LSU Neuroscience Center, Louisiana State University Health Sciences Center, New Orleans, Louisiana 70112-2272, USA.
Zhao
Yuhai
Y
Lukiw
Walter J
WJ
D000368:Aged; D001921:Brain / Q000502:physiology*; D001923:Brain Chemistry / Q000235:genetics*; D002467:Cell Nucleus / Q000502:physiology; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification*; D014158:Transcription, Genetic
D012333:RNA, Messenger