Updated : Thu, June 29, 2023
Whether you’re working with samples for PCR or sequencing, it's worth questioning what type of extraction or purification method would work well with your downstream application. Are there potential time and cost efficiencies? Do you need ultra-pure nucleic acid, or will a simple crude prep do the job just as well?
There are a few things to consider when it comes to deciding which method of extraction and purification are the most appropriate:
- How challenging is your sample type?
- Is it full of contaminants that may impact your downstream applications?
- What are your requirements for throughput/speed, simplicity and automation?
- Do you need high yields and/or high-quality nucleic acids?
- Do you want to avoid working with hazardous extraction reagents?
For many sample types and downstream applications such as PCR and RT-PCR, extracting nucleic acids without further purification steps is often all that’s required. For other applications, as well as particular sample types, pairing a rapid and efficient extraction method, such as LGC Biosearch Technologies’ QuickExtract™ DNA and RNA solutions, with a follow up purification technique where necessary can be a powerful technique to get the highest quality nucleic acids that you need, while saving you time and effort.
So is it really feasible to ditch the spin columns? Well, if you are considering making the switch to a simpler, faster and more cost-effective method then you’re not alone.
Sample prep that’s out of this world
Sarah Stahl-Rommel and her colleagues at NASA1 switched from carrying out microbial monitoring of the International Space Station (ISS) by culture-dependent methods requiring samples to be returned to the ground to a culture-independent, swab-to-sequencer method. This change, enabled by QuickExtract, provided astronauts with the autonomy to generate real-time data, using non-toxic reagents and a method simple enough for non-specialists to carry out onboard the ISS with limited equipment. 
Cutting out the extraction step for SHERLOCK one-pot testing
SHERLOCK methods, usually performed with an initial target amplification step followed by CRISPR-mediated nucleic acid detection, have been used in diagnostic tests for SARS-CoV-2. Joung et al.2 streamlined this process by using QuickExtract and cutting out the separate RNA extraction step. This simplified assay can be performed as a one-pot test, combining RNA extraction with isothermal amplification and CRISPR-mediated detection, making it more suitable for point of care applications.
Boosting the efficiency of gene editing
Ghasemi et al.3 described a method to boost the efficiency of gene editing via homology-directed repair. They achieved this by covalently modifying the template DNA with interstrand crosslinks, increasing Cas9-mediated editing efficiencies by up to fivefold. The authors of this study used QuickExtract to prepare genomic DNA prior to amplicon sequencing.
Quick and easy prep for drag and drop genome insertion
Yarnall et al.4 created a new method to integrate large, diverse DNA cargo into genomes without relying on DNA repair pathways. Enabled by QuickExtract for quick and easy genomic DNA extraction, programmable addition via site-specific targeting elements (PASTE) uses CRISPR-Cas9 nickase, fused to a reverse transcriptase and serine integrase. Results showed that sequences as large as ~36 kilobases could be integrated using this method.
Simple and fast extraction for cancer immunotherapy research
Treatment strategies to effectively overcome resistance to cancer immunotherapy are sadly lacking. Sun et al.5 recently identified a new immune-evasion gene, TBK1, that can be targeted to help overcome resistance. QuickExtract was used in their work for simple and fast DNA extraction prior to PCR targeting the Tbk1 gene.
Ultra-rapid detection of somatic variants via real-time targeted amplicon sequencing
Sequencing-based detection of molecular markers for cancer detection is considered the gold standard method, but usually is time consuming due to the laborious sample preparation steps. Wadden et al.6 presented a new approach, using LAMP and a bioinformatics tool, leading to a result in less than 30 minutes. They used QuickExtract for a rapid, one-pot protocol resulting in DNA that was immediately suitable for input for PCR amplification if diluted or LAMP.
Bypassing the RNA isolation step in the
point of care diagnostics market
RT-qPCR-based diagnostic tests play a huge role in the point of care diagnostics market, but often come with high associated costs which limit their accessibility. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is an alternative detection method that can be carried out with basic equipment at lower costs. Kellner et al.7 developed a rapid 40-minute procedure for home testing that manages to skip the RNA isolation step by using QuickExtract to heat-inactivate crude lysate. They demonstrated that this performed equally well compared to a standard RNA extraction step, while saving time and expense.
So how does QuickExtract work?
With the entire process taking place within a single tube, there is no need for centrifugation or spin columns. The superfast extraction protocol can be done in 3-8 minutes, using only non-toxic reagents, resulting in PCR-grade nucleic acid from almost any sample type.
Talk to us
Choosing an appropriate method of nucleic acid extraction and/or purification can be critical for your workflow. At Biosearch Technologies, our expert scientists can advise you on which isolation method best suits your needs and can offer flexible custom and bulk solutions. Get in touch today.
Related content
Blog post: How to select an effective nucleic acid isolation method for human diagnostics?
Application note: (created in partnership with SelectScience) Choose an optimal extraction or purification kit best suited to your sample type and downstream requirements.
Application note: See how QuickExtract is compatible with various human pathogen diagnostic samples.
References
- Stahl-Rommel S, Jain M, Nguyen HN, et al. (2021). Real-Time Culture-Independent Microbial Profiling Onboard the International Space Station Using Nanopore Sequencing. Genes. 12(1):106. DOI:10.3390/genes12010106
- Joung J, Ladha A, Saito M, et al. (2020). N Engl J Med, 383:1492-1494. DOI:10.1056/NEJMc2026172
- Ghasemi HI, Bacal J, Yoon AC, et al. (2023). Interstrand crosslinking of homologous repair template DNA enhances gene editing in human cells. Nat Biotechnol. DOI:10.1038/s41587-022-01654-y
- Yarnall MTN, Ioannidi EI, Schmitt-Ulms C, et al. (2023). Drag-and-drop genome insertion of large sequences without double-strand DNA cleavage using CRISPR-directed integrases. Nat Biotechnol 41, 500–512. DOI:10.1038/s41587-022-01527-4
- Sun Y, Revach Oy, Anderson S. et al. (2023). Targeting TBK1 to overcome resistance to cancer immunotherapy. Nature 615, 158–167. DOI:10.1038/s41586-023-05704-6
- Wadden J, Newell BS, Bugbee J, et al. (2022). Ultra-rapid somatic variant detection via real-time targeted amplicon sequencing. Commun Biol 5, 708. DOI:10.1038/s42003-022-03657-6
- Kellner MJ, Ross JJ, Schnabl J, et al. (2022). A Rapid, Highly Sensitive and Open-Access SARS-CoV-2 Detection Assay for Laboratory and Home Testing. Front Mol Biosci. 9:801309. DOI:10.3389/fmolb.2022.801309
