Experiments in the preparation of frozen tissues and sections
For better results, fresh tissue should be used, because samples stored at -80°C for more than 24 h will have much lower RNA yield and integrity. Although seve
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Competitive RT-PCR: Evaluation of copy number
In the preexperiment, reverse transcription should be done separately for the sample and the competitor, and then the unchanged amount of this sample reverse tr
HE staining of slices for LCM
Prior to LCM, cut tissue samples are fixed, stained, and dehydrated if necessary. Authors use classical HE staining for visceral and lymph node tissues. Once th
Competitive RT-PCR: Competitor RNA construction assay
Part I: Design of competitor; Part II: Synthesis, purification and quantification of competitor RNA. This experiment is from PCR Laboratory Guide (Second Editio
On-column removal assay for DNA contamination
This protocol can be used for RNA isolation strategies using silica-based RNA binding columns, such as protocol 6, however, 100% DNA removal may not be achieved
Relative RT-PCR: Determination of Primer-to-Competitor Ratio Experiments
The purpose of this scheme is to select an appropriate ratio between the 18SrRNA primer and the competitor that will mimic the expression of the target transcri
Single-stranded cDNA synthesis
This protocol utilizes total RNA because if 18SrRNA is used as an internal control, RNA with poly(A) cannot be used in subsequent experiments. and the RNA must
RNA extraction experiments from a limited number of cells
RNA samples can be stored at -80°C for up to 6 months. Total cellular RNA is extracted using the StratageneRNA Extraction Kit, following the instructions. Alte
Relative RT-PCR: Sample quantification assay
Knowing the linear range of cycling parameters and the ratio of 18SrRNA primer to competitor, one can use one's own samples for relative quantitative RT-PCR.The