In selecting an RNA isolation kit, not only must the ability of the kit to recover the required amount of total RNA be taken into consideration, but also the ability of the kit to recover RNA meeting specific quality criteria.
An examination of the %CV for the amount of RNA recovered by each kit showed that the Plus Universal Kit and Maxwell® kits were the most consistent in their ability to isolate RNA (%CV =6.9 and 16.1, respectively).
Total RNA was extracted from formalin-fixed paraffin-embedded (FFPE) tissue using Recover All kit (Ambion, Carlsbad, CA, USA).
The Plus Universal kit consistently produced RNA samples of sufficient quantity; however, none of the samples extracted with this kit had RIN values ≥7 suggesting that the RNA recovered via this kit is subject to fragmentation and may be less than optimal for microarray and NGS applications.
Despite significant differences in the quantity of RNA recovered by each kit, the amount of RNA recovered from each tissue sample was ≥1 μg, independent of the kit used - with the exception of blood extracted via the SimplyRNA HT kit (average yield of 1.74 ± 1.7 μg).
For Northern blots, RNA was prepared from bacteria stored in RNAprotect Bacteria Reagent and recovered from RNeasy kit spin columns according to the manufacturer's protocols (Qiagen, Valencia, CA).
Although the CNA kit recovered a high percentage (83%%) of the smallest plasmid fragment (115 bp), the yield of this was not as high as that of the 461-bp fragment (99%%) (p < 0.05).
With regard to variation in the amount of total RNA recovered by each kit, no significant differences in variance were detected, with the exception of embryos, indicating that the consistencies of each kit with regard to the amount of RNA extracted were similar.
As part of the process to help children recover, volunteers use UNICEF recreation kits containing books, crayons, games, toys, musical instruments and sporting equipment.
Several manufacturers have recently begun to market kits designed to recover high-quality RNA from these types of samples [ 10]; however, few studies have directly compared the performance of commercially available kits in isolating RNA from small tissue samples [ 11].
A closer examination of the quality of RNA extracted using each of these kits, revealed that the Plus Mini, Plus Universal and Maxwell® kits recovered RNA with A260 A280 ratios ≥1.8 and RIN ≥5 from blood, spleen, kidney and larvae indicating that RNA extracted from these tissues via these kits is acceptable for qPCR.
