By John R. Gosden
Innovative researchers reveal step by step how oligonucleotide primers could be effectively used to become aware of and magnify or expand complimentary sequences in situ. via those procedures-often invented by means of the authors-the door is opened to swift identity and characterization of chromosomal DNA sequences, viral genomes, and infrequent messenger RNAs in cells-at hitherto unequalled levels of sensitivity and specificity. Their leading edge techniques-suitable for either amateur and skilled researchers-have quickly develop into crucial for plenty of medical diagnostic strategies, even if in quantification of chromosomes within the id of aneuploidy for prenatal prognosis, or within the id of viral an infection within the early phases, or of infrequent messenger RNAs found in cells.
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Extra resources for PRINS and In Situ PCR Protocols
1995) Cytochemical detection systems for in situ hybridization, and the combination with mnnunocytochemistry. Histochem. J. 21,833-858. 10. Speel, E. J. , Ramaekers, F. C. , and Hopman, A. H. N. (1994) Combined immunocytochemistry and fluorescence in situ hybridization for simultaneous tricolor detection of cell cycle, genomic, and phenotypic parameters of tumor cells. J. Histochem. Cytochem. 42,96 1-966. 11. , Lewis, W. , and Jones, C. (1989) A finestructure deletion map of chromosome 1 lp: analysis of Jl series hybrids.
PRINS hybridization developed by Koch et al. (4) was a clever improvisation using oligonucleotide primers to anneal to chromosomal DNA sequencesfollowed by extension using DNA polymerase. A reporter molecule (a digoxigenin or biotinylated deoxynucleotide triphosphate) was incorporated during the reaction. The reporter molecule was then detected using an immunocytochemical approach. Improvements to the basic PRINS technique involve direct incorporation of fluorochrome-tagged deoxynucleotide triphosphates, negating the need for secondary labeling and, the “cycling PRINS” (5) reactions.
And Litt, G J. (1989) Catalyzed reporter deposition, a novel method of signal amplification. Amplification to immunoassays. J. Immunol. Methods 125,279-285. 13. Speel, E. J. , Ramaekers, F. C. , and Hopman, A. H. N. (1995) Cytochemrcal detection systems for in situ hybridrzation, and the combmatron with immunocytochemistry. Histochem. J 27,833-858. Multiple Sequential Oligonucleotide Primed /n Sctu DNA Syntheses (MULTI-PRINS) John R. Gosden and Diane Lawson 1. Introduction Conventional PRINS (if it is possible to use such a description for a relatively new technique) is capable of identifying and quantifying chromosomes or chromosome pairs in metaphase or interphase cells (1-4).