Chapter 5
Introduction to the Detection of Specific
DNA and RNA Sequences
TAKEHIKO KOJI
Historical remarks
In situ hybridization was originally developed by Gall and Pardue (1969) and John et al. (1969) as a method to localize specific
DNA sequences in cytological preparation. Afterwards, the
method was used as a powerful technique to detect the locus
of a specific gene in chromosomes or the sites of viral DNA.
In the late 1970s, Brahic and Haase (1978) revealed that the
same principle is applicable to localize specific RNA in cell
and tissue preparations. In keeping pace with the burst prosperity of molecular cloning fields, in situ hybridization has progressed and now has turned out to be one of the most essential
techniques in cell biology and developmental biology. In this
chapter, I will try to overview the methodological aspects of
in situ hybridization, particularly focusing on the detection of
specific RNA in tissue sections, in which we have been actively
engaged for more than 15 years (Koji and Nakane 1990, 1996).
For a better understanding of the regulatory mechanisms of
cell-type or tissue-type specific gene expression, it is essential to
analyze the expression of specific genes at the level of mRNA as
well as protein in individual cells. To localize proteins at a cellular level, immunohistochemistry has been a powerful tool.
However, the information obtained by immunohistochemistry
on intracellular protein synthesis was limited because the protein might be secreted, degradated, or post-translationally modified. Also, it was known that the best performance of immunosCorrespondence to Takehiko Koji, Nagasaki University School of Medicine,
Department of Histology and Cell Biology, 1-12-4 Sakamoto, Nagasaki, 8528523, Japan (phone +81-095-849-7025; fax +81-095-849-7028; e-mail tkoji@net.nagasaki-u.ac.jp)
Introduction to the Detection of Specific
DNA and RNA Sequences
TAKEHIKO KOJI
Historical remarks
In situ hybridization was originally developed by Gall and Pardue (1969) and John et al. (1969) as a method to localize specific
DNA sequences in cytological preparation. Afterwards, the
method was used as a powerful technique to detect the locus
of a specific gene in chromosomes or the sites of viral DNA.
In the late 1970s, Brahic and Haase (1978) revealed that the
same principle is applicable to localize specific RNA in cell
and tissue preparations. In keeping pace with the burst prosperity of molecular cloning fields, in situ hybridization has progressed and now has turned out to be one of the most essential
techniques in cell biology and developmental biology. In this
chapter, I will try to overview the methodological aspects of
in situ hybridization, particularly focusing on the detection of
specific RNA in tissue sections, in which we have been actively
engaged for more than 15 years (Koji and Nakane 1990, 1996).
For a better understanding of the regulatory mechanisms of
cell-type or tissue-type specific gene expression, it is essential to
analyze the expression of specific genes at the level of mRNA as
well as protein in individual cells. To localize proteins at a cellular level, immunohistochemistry has been a powerful tool.
However, the information obtained by immunohistochemistry
on intracellular protein synthesis was limited because the protein might be secreted, degradated, or post-translationally modified. Also, it was known that the best performance of immunosCorrespondence to Takehiko Koji, Nagasaki University School of Medicine,
Department of Histology and Cell Biology, 1-12-4 Sakamoto, Nagasaki, 8528523, Japan (phone +81-095-849-7025; fax +81-095-849-7028; e-mail tkoji@net.nagasaki-u.ac.jp)