The isolation and in vitro translation of undegraded messenger RNAs from human postmortem brain.

Author(s): Morrison MR, Griffin WS

Publication: Anal Biochem, 1981, Vol. 113, Page 318-24

PubMed ID: 7283137 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to investigate the effects of PMI (4 h, or 16 h), biospecimen storage (fresh, or snap-frozen) and RNA isolation method on RNA yield and in vitro translation products from cerebella. Each PMI in this study was represented by a single individual.

   Summary of Findings:

   A PMI of 16 h resulted in an RNA yield of only 70% of that if the PMI was 4 h. Correspondingly, less protein was produced by in vitro translation after a PMI of 16 h than 4 h. Two dimensional gel analysis of the in vitro translation products did not reveal any effect of PMI or specimen processing on relative transcript distribution, but less overall signal was observed. The use of fresh versus snap-frozen specimens and inclusion of vanadium ribonucleoside complex in the aqueous extraction buffer did not alter RNA yield or in vitro translation regardless of PMI. RNA yield for specimens with a PMI of 4 h and 16 h was increased 1.65 fold if the specimens were homogenized in guanidium chloride versus the aqueous extraction buffer, but no change in the size distribution of in vitro translation products was observed. The authors conclude that a PMI of 16 h results in less RNA yield, but the effect is not due to degradation of specific transcripts but rather general degradation of all transcripts.

   Biospecimens

   • Tissue - Brain

   Preservative Types

   • None (Fresh)
   • Frozen

   Diagnoses:

   • Autopsy

   Platform:

   Analyte	Technology Platform
   RNA	Spectrophotometry
   RNA	In vitro translation

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Preaquisition	Postmortem interval	4 h 16 h
   Biospecimen Preservation	Type of fixation/preservation	Snap frozen None (fresh)
   Analyte Extraction and Purification	Analyte isolation method	Homogenization in aqueous buffer with vanadium ribonucleoside complex Homogenization in aqueous buffer without vanadium ribonucleoside complex Homogenization in guanidium chloride

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