( which involves tissue/cell l...)Robust High Throughput RNA Isolation from Blood Samples

which involves tissue/cell lysis and magnetic bead sample capture followed by TURBO DNase treatment. High quality, intact RNA with 28S/18S rRNA ratios >0.9 (Agilent 2100 bioanalyzer) was obtained (Figure 1). RNA yield was determined by A260 with an average range of 0.310.52 g total RNA from all samples (50 l each).

Figure 1. MagMAX-96 Blood Consistently Recovers Intact Total RNA from Fresh Blood and Blood Stored at 4C. Human whole blood from four different donors was collected into EDTAVacutainer tubes; aliquots were stored at 4C for 24 hr. Fresh whole blood was collected from the same donors the following day for concurrent RNA isolation with the samples stored at 4C. Total RNA was isolated from 50 l blood from multiple donors in 8 replicates using the MagMAX-96 Blood Kit, and total RNA (~50 ng) from representative samples was analyzed on an Agilent 2100 bioanalyzer (A). RNA yield was determined by A260 measurement using the NanoDrop ND-1000A Spectrophotometer (B).

Genomic DNA contamination was determined by one-step qRT-PCR targeting hTBP (human TATA binding protein) and RPII (RNA polymerase II) mRNAs [(+)RT reactions] and genes [(-)RT reactions]. Total RNA purified from all samples contained minimal genomic DNA contamination as assessed by delta-Ct values from qRT-PCR (Figure 2). Similar (+)RT Ct values of qRT-PCR (hTBP and RPII) using RNA isolated from fresh blood and blood stored at 4C were obtained.

Figure 2. Comparison of Total RNA Isolated from Fresh Blood and Blood Stored at 4C in qRT-PCR A
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