ArticleActa histochemica et cytochemica2009
Improvement of in situ PCR by optimization of PCR cycle number and proteinase k concentration: localization of x chromosome-linked phosphoglycerate kinase-1 gene in mouse reproductive organs.
Article in Acta histochemica et cytochemica, 2009. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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7 citing papers in PubMed, 10 citations in OpenAlex.
- Global changes in epigenomes during mouse spermatogenesis: possible relation to germ cell apoptosis.Histochemistry and cell biology · 2020Review
- Detecting the colonization of ericoid mycorrhizal fungi inPlant methods · 2020Article
- Localisation of abundant and organ-specific genes expressed in Rosa hybrida leaves and flower buds by direct in situ RT-PCR.TheScientificWorldJournal · 2012Article
- Immunohistochemical Analysis of Histone H3 Modifications in Germ Cells during Mouse Spermatogenesis.Acta histochemica et cytochemica · 2011Article
- Animal models of middle ear cholesteatoma.Journal of biomedicine & biotechnology · 2011Review
- Pathogenesis of middle ear cholesteatoma: a new model of experimentally induced cholesteatoma in Mongolian gerbils.The American journal of pathology · 2010Article
- Human papillomavirus infection and its possible correlation with p63 expression in cervical cancer in Japan, Mongolia, and Myanmar.Acta histochemica et cytochemica · 2009Article
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5 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
In situ polymerase chain reaction (in situ PCR), which can detect a few copies of genes within a cell by amplifying the target gene, was developed to better understand the biological functions of tissues. In this study, we optimized the protocol conditions for the detection of X chromosome-linked phosphoglycerate kinase-1 (pgk-1) gene in paraffin-embedded sections of mouse reproductive organs. The effects of various concentrations of proteinase K (PK) and PCR cycle numbers were examined. To label the amplified DNA, we used digoxigenin-dUTP (Dig), Cy-3-dUTP (Cy-3), or FluorX-dCTP (FluorX). The optimal concentration of PK was 50 microg/ml for the ovary and 10 microg/ml for the testis. Ten PCR cycles were optimal for Dig and 25 cycles were optimal for FluorX and Cy-3 in the ovary and testis. The signal-to-noise ratio of FluorX and Cy-3 for ovarian tissue was better than that of Dig. Using the above conditions, we detected 1-4 and 1-2 spots of pgk-1 in the nuclei of granulosa and germ cells, respectively. Our results indicate that in situ PCR is useful for detecting a specific gene in paraffin-embedded sections under optimized conditions of both PCR cycle number and PK concentration.
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