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New Kit Generates Exceptionally Pure Total RNA for RT-PCR

ion factor 1a), medium-abundance transcripts (b2-microglobulin), and low-abundance transcripts (protein phosphatase 1 and ornithine decarboxylase). Although b2-microglobulin mRNA is classified as medium abundance, its expression level is higher in liver than in brain7 (Figure 3). Similarly, ornithine decarboxylase is expressed at overall low levels but the levels vary in tissues throughout the body.8 The presence of the expected PCR bands in Figure 3 shows that even rare RNA transcripts are represented in total RNA isolated with the Absolutely RNA kit.

Conclusions

RNA isolated with Stratagenes Absolutely RNA RT-PCR miniprep kit is ideal for RT-PCR. The kits quick, convenient protocol includes a DNA removal step that ensures sensitive RT-PCR results without having amplification products generated from genomic DNA targets. Moreover, RT-PCR readily detects low-abundance transcripts from this RNA. The Absolutely RNA RT-PCR miniprep kit effectively generates high-purity, intact total RNA from a range of tissues and cultured cells, ready for the most sensitive RT-PCR experiments.2

Methods

The ProSTAR first strand RT-PCR kit was used for cDNA synthesis.3 For GAPDH primer detection of DNA in total RNA samples, control cDNA synthesis reactions were performed in the absence of reverse transcriptase. PCR reactions contained 1 l from the cDNA reactions (100 ng of total RNA), 200 M of each dNTP, 25 ng of each GAPDH primer, 1X Taq polymerase buffer, and 1.25 U of Taq2000 DNA polymerase. PCR react
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