Semi-Automation Drives RNA Purification in Various Cells and Tissues

One of the simplest ways to increase both speed and accuracy in the laboratory is to exploit the benefits of semi-automation. By limiting hands-on requirements, research teams can improve overall efficiency while ensuring clean results.

The KingFisher Pure RNA Tissue Kit, in combination with either the KingFisher Flex or Duo instruments (all Thermo Scientific), is one means of semi-automating RNA purification. The kit itself is suitable for purifying RNA from mammalian, yeast and bacterial samples. Since RNA preservation is a primary concern at every step of the process—from collection to storage to purification—the manufacturer specifically designed the kit to preserve RNA in the interest of optimized yield and quality. In terms of throughput, the Flex and Duo platforms can automate 96 samples or 12 samples, respectively, per 60-minute run.

Of course, while ease of use and efficiency are key considerations, yield and quality remain paramount. The SP&A Application Laboratory¹ evaluated these very things using 12 sample types: three mammalian cell lines, seven mouse tissues, one bacterial sample and one yeast culture. The bacterial and yeast samples required additional reagents for lysis. After purification, total RNA eluted into 100 μL nuclease-free water, although elution volume is adjustable using BindIt software (Thermo Scientific).

The team ran the total RNA eluates from all 12 samples on 1% agarose gel followed by Agilent Bioanalyzer RNA 6000 Nano Chip. They reported two clear bands and high RNA integrity number (RIN) values, assuring both high yield and excellent product quality. The team verified reproducibility using culture HeLa cells with consistent results. The RIN values also showed good between-well reproducibility.

Overall, the laboratory found the kit to meet its standards for efficiency and ease of use while providing sufficient yield of high-integrity RNA over a wide range of sample types. They note that the kit components sweep away contaminants and inhibitors, producing eluate appropriate for direct use in downstream applications. This system is one option for maintaining critical RNA preservation along the entire chain of custody.

RIN values from total RNA from various sample types¹

Sample	Starting volume	RIN
Jurkat cells	1 × 106 cells	8.1
HeLa cells	1 × 106 cells	9.8
Cos-7 cells	5 × 105 cells	10
Mouse heart	5 mg	8.9
Mouse liver	5 mg	7.8
Mouse brain	5 mg	8.4
Mouse lung	5 mg	7.2
Mouse muscle	5 mg	7.2
Mouse kidney	5 mg	8.5
Mouse spleen	5 mg	9.2
E. coli ER2267	1 mL overnight culture	9.4
Saccharomyces cerevisiae	1 mL overnight culture	N/A

 

Reference

1 SP&A Application Laboratory. (2013) “Purification of high-quality RNA from cell and tissue samples with the Thermo Scientific KingFisher Pure RNA Tissue Kit,” Thermo Fisher Scientific.