Method for quantifying total mRNA with poly(A)-complementary oli

Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid

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536 243, 536 2532, 4352872, 4352877, 4352883, 4352884, 424 92, 4241811, C12Q 168

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059767979

ABSTRACT:
A method for quantifying total mRNA in a biological sample containing RNA such as crude cell lysates containing cytosolic mRNA, which method comprises the steps of: (a) incubating the sample with an oligo-(dT)- or poly-U-immobilized microtiter plate; (b) washing non-hybridized components from the microtiter plate; (c) labeling the hybridized mRNA with a photometric nucleic-acid dye; (d) measuring the amount of label captured on the microtiter plate; (e) heat-denaturing the labeled mRNA; (f) washing the denatured mRNA from the microtiter plate; and (g) measuring the amount of label remaining on the microtiter plate; and (h) correlating the amount of the measured label (captured label minus remaining label) with the quantity of total mRNA present in the sample, thereby easily measuring the total mRNA without the influence of rRNA or tRNA and without radioactive dyes, which method can be adapted to chemosensitivity tests.

REFERENCES:
Shroder et al. Biochemistry 29: 2368-2378, Sep. 1990.

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