Amplified RNA synthesized from limited quantities of heterogeneous cDNA.

DOI Web Site 被引用文献23件
  • R N Van Gelder
    Department of Psychiatry, Stanford University School of Medicine, CA 94305.
  • M E von Zastrow
    Department of Psychiatry, Stanford University School of Medicine, CA 94305.
  • A Yool
    Department of Psychiatry, Stanford University School of Medicine, CA 94305.
  • W C Dement
    Department of Psychiatry, Stanford University School of Medicine, CA 94305.
  • J D Barchas
    Department of Psychiatry, Stanford University School of Medicine, CA 94305.
  • J H Eberwine
    Department of Psychiatry, Stanford University School of Medicine, CA 94305.

書誌事項

公開日
1990-03
DOI
  • 10.1073/pnas.87.5.1663
公開者
Proceedings of the National Academy of Sciences

この論文をさがす

説明

<jats:p>The heterogeneity of neural gene expression and the spatially limited expression of many low-abundance messenger RNAs in the brain has made cloning and analysis of such messages difficult. To generate amounts of nucleic acids sufficient for use in standard cloning strategies, we have devised a method for producing amplified heterogeneous populations of RNA from limited quantities of cDNA. Whole cerebellar RNA was primed with a synthetic oligonucleotide containing the T7 RNA polymerase promoter sequence 5' to a polythymidylate region. After second-strand cDNA synthesis, T7 RNA polymerase was used to generate amplified antisense RNA (aRNA). Up to 80-fold molar amplification has been achieved from nanogram quantities of cDNA. The amplified material is similar in size distribution to the parent cDNA and shows sequence heterogeneity as assessed by Southern and Northern blot analysis. Specific messages for moderate-abundance mRNAs for actin and guanine nucleotide-binding protein (G-protein) alpha subunits have been detected in the amplified material. By using in situ transcription to generate cDNA, sequences for cyclophilin have been detected in aRNA derived from single cerebellar tissue sections. cDNA derived from a single cerebellar Purkinje cell also has been amplified and yields material that hybridizes to cognate whole RNA and mRNA but not to Escherichia coli RNA.</jats:p>

収録刊行物

被引用文献 (23)*注記

もっと見る

詳細情報 詳細情報について

問題の指摘

ページトップへ