Assessment of normalization strategies for quantitative RT-PCR using microdissected tissue samples
نویسندگان
چکیده
منابع مشابه
One-step RT-PCR for screening microdissected tissue samples.
The technology of laser-capture microdissection (LCM) enables the easy extraction of single cells or defined groups of cells from a tissue section (1,2). This technique uses a laser pulse to activate a plastic polymer, which binds to and captures cells from a frozen or paraffin-embedded tissue section. Captured cells present on this solid support can then be used for nucleic acid and/or protein...
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This chapter describes a method for the rapid assessment of gene copy number in laser microdissected material using multiplex real-time polymerase chain reaction (PCR). Here a putative oncogene (ZNF217) was evaluated in a series of colon tumors, but the method is applicable to any locus for which a nucleic acid sequence is available. The preparation, laser microdissection, and optimum storage o...
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Quantitative RT-PCR is often used as a research tool directed at gene transcription. Selection of optimal housekeeping genes (HKGs) as reference genes is critical to establishing sensitive and reproducible qRT-PCR-based assays. The current study was designed to identify the appropriate reference genes in blood leukocytes of bottlenose dolphins (Tursiops truncatus) for gene transcription researc...
متن کاملOne-step RT-PCR without initial RNA isolation sStep for laser-microdissected tissue sample.
One-step RT-PCR procedure without initial RNA extraction step is tested for laser microdissected tissue sample. Unfixed cryosections of liver and kidney tissue of male SD rats were cut using laser microdissection system and directly used as templates for RT-PCR study. To check the sensitivity, 5, 25, 125, and 625 hepatocytes were cut and put in PCR-tube. After DNase treatment and cDNA synthesis...
متن کاملGuidelines for quantitative rt-PCR.
Something I notice often in supervising new graduate students is their frustration with the irreproducibility of their biological data. For example, I am asked frequently ‘‘how many biological replicates do I need for qRT-PCR’’? The answer, of course, lies first with the students themselves. They need as many replicates as will persuade them of the validity of their observations. However, for m...
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ژورنال
عنوان ژورنال: Laboratory Investigation
سال: 2007
ISSN: 0023-6837,1530-0307
DOI: 10.1038/labinvest.3700659