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In Situ Hybridization for RNA: Nonradioactive Probe: ss cDNA Probe

  • Yoshio Kanemitsu
  • Takehiko Koji
Chapter
Part of the Springer Lab Manuals book series (SLM)

Abstract

The use of single-stranded cD NA (ss cDNA) is more advantageous in sensitivity than that of double-stranded cD NA (ds cDNA) or synthetic oligonucleotide as a probe for in situ hybridization (ISH) (Sugawara et al. 1990). In this chapter, we present a simple procedure for the production of nonradioactive ss cD NA probe by combining the two-step asymmetric PCR and thyminse-thyminse dimer (T-T dimer) haptenization.

Keywords

Thermal Controller Vortex Briefly Nonradioactive Probe Optimum Time Parameter Versus Reverse Transcriptase 
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References

  1. Chomczynski P and Sacchi N (1987) Single-step method of RNA isolation by acid guanidium thiocyanate-phenol-chloroform extraction. Analytical Biochemistry 162: 156–159PubMedCrossRefGoogle Scholar
  2. Kanemitsu Y Koji T Hori T and Nakane P.K. (1994) Production of anti sense DNA probe by asymmetric PCR and its application for in situ hybridization. Nucleic Acids Symposium Series 31: 57–59Google Scholar
  3. Koji T and Nakane P.K. (1990) Localization in situ of specific mRNA using thymine-thymine dimerized DNA probes. Sensitive and reliable nonradioactive in situ hybridization. Acta Pathol Jpn 40: 793–807PubMedGoogle Scholar
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Copyright information

© Springer Japan 2000

Authors and Affiliations

  • Yoshio Kanemitsu
    • 1
  • Takehiko Koji
    • 2
  1. 1.Department of Physiology, Faculty of MedicineKyushu UniversityFukuokaJapan
  2. 2.Department of AnatomyNagasaki University School of MedicineNagasakiJapan

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