DNA sequesters endogenous mRNA during preparation of crude Escherichia coli extracts for protein synthesis; use of an S60 reduces the sequestered mRNA

Emanuel Goldman; Judith Gibel

doi:https://doi.org/10.1016/0003-9861(83)90197-2

DNA sequesters endogenous mRNA during preparation of crude Escherichia coli extracts for protein synthesis; use of an S60 reduces the sequestered mRNA

Emanuel Goldman, Judith Gibel

NJMS-Molecular Core

Rutgers, The State University

Research output: Contribution to journal › Article › peer-review

2 Scopus citations

Abstract

High backgrounds of endogenous incorporation of amino acids into protein in crude extracts of Escherichia coli are a consequence of endogenous messenger RNA. This RNA survives the preparation by virtue of protection or sequestering by endogenous DNA. Thus, DNase treatment in the crude extract leads to the elimination of this mRNA, while DNase treatment has no effect on the purified RNA. The endogenous mRNA also appears to be physically associated with DNA on CsCl gradients, and can be largely removed along with DNA by centrifugation of extracts at 60,000g. The top layer above a 60,000g centrifugation (S60) appears to be suitable for protein synthesis, and provides for lower background levels of endogenous messenger RNA.

Original language	English (US)
Pages (from-to)	134-141
Number of pages	8
Journal	Archives of Biochemistry and Biophysics
Volume	224
Issue number	1
DOIs	https://doi.org/10.1016/0003-9861(83)90197-2
State	Published - Jul 1 1983

ASJC Scopus subject areas

Biophysics
Biochemistry
Molecular Biology

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https://doi.org/10.1016/0003-9861(83)90197-2

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title = "DNA sequesters endogenous mRNA during preparation of crude Escherichia coli extracts for protein synthesis; use of an S60 reduces the sequestered mRNA",

abstract = "High backgrounds of endogenous incorporation of amino acids into protein in crude extracts of Escherichia coli are a consequence of endogenous messenger RNA. This RNA survives the preparation by virtue of protection or sequestering by endogenous DNA. Thus, DNase treatment in the crude extract leads to the elimination of this mRNA, while DNase treatment has no effect on the purified RNA. The endogenous mRNA also appears to be physically associated with DNA on CsCl gradients, and can be largely removed along with DNA by centrifugation of extracts at 60,000g. The top layer above a 60,000g centrifugation (S60) appears to be suitable for protein synthesis, and provides for lower background levels of endogenous messenger RNA.",

author = "Emanuel Goldman and Judith Gibel",

note = "Funding Information: This work was supported by a grant from the National Institutes of Health (GM27{\textquoteright}711). E.G. is a recipient of a Research Career Development Award from the National Cancer Institute (CA 06890).",

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T1 - DNA sequesters endogenous mRNA during preparation of crude Escherichia coli extracts for protein synthesis; use of an S60 reduces the sequestered mRNA

AU - Goldman, Emanuel

AU - Gibel, Judith

N1 - Funding Information: This work was supported by a grant from the National Institutes of Health (GM27’711). E.G. is a recipient of a Research Career Development Award from the National Cancer Institute (CA 06890).

PY - 1983/7/1

Y1 - 1983/7/1

N2 - High backgrounds of endogenous incorporation of amino acids into protein in crude extracts of Escherichia coli are a consequence of endogenous messenger RNA. This RNA survives the preparation by virtue of protection or sequestering by endogenous DNA. Thus, DNase treatment in the crude extract leads to the elimination of this mRNA, while DNase treatment has no effect on the purified RNA. The endogenous mRNA also appears to be physically associated with DNA on CsCl gradients, and can be largely removed along with DNA by centrifugation of extracts at 60,000g. The top layer above a 60,000g centrifugation (S60) appears to be suitable for protein synthesis, and provides for lower background levels of endogenous messenger RNA.

AB - High backgrounds of endogenous incorporation of amino acids into protein in crude extracts of Escherichia coli are a consequence of endogenous messenger RNA. This RNA survives the preparation by virtue of protection or sequestering by endogenous DNA. Thus, DNase treatment in the crude extract leads to the elimination of this mRNA, while DNase treatment has no effect on the purified RNA. The endogenous mRNA also appears to be physically associated with DNA on CsCl gradients, and can be largely removed along with DNA by centrifugation of extracts at 60,000g. The top layer above a 60,000g centrifugation (S60) appears to be suitable for protein synthesis, and provides for lower background levels of endogenous messenger RNA.

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DNA sequesters endogenous mRNA during preparation of crude Escherichia coli extracts for protein synthesis; use of an S60 reduces the sequestered mRNA

Abstract

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