How to increase the quality and yield of DNA and RNA from saliva

How to increase the quality and yield of DNA and RNA from saliva


With advanced genomic analysis applications becoming routine in many labs, the need for high purity, high quality DNA and RNA samples collected using non-invasive sampling methods, such as saliva, is becoming increasingly important.

Nucleic acid extraction from saliva can be challenging due to the presence of contaminants and inhibitors  that can interfere with downstream applications. However, in recent years significant improvements have been made with devices to collect, store, and process saliva to extract DNA and RNA. Good yields of high-quality nucleic acids can now be extracted for use in sensitive downstream applications.

Variations in sample collection and handling can dramatically affect data quality. This article explains how to get the best from your saliva samples by maximizing the purity and yields of DNA and RNA with information on:

  • Preventing sample contamination
  • Saliva collection from donors
  • Nucleic acid stabilization
  • Safely shipping saliva samples
  • Choosing your DNA/RNA isolation kit
  • Isolating high molecular weight (HMW) DNA from saliva   
  • Cleaning up your DNA after an initial extraction

Preventing sample contamination

The main sources of human DNA in saliva are epithelial cells and leukocytes. Saliva samples may also include bacterial DNA, allowing for DNA extraction and analysis of the oral microbiome. To prevent contamination from external nucleic acids and other factors that may interfere with the purity of samples, it is important to ensure that donors do not eat, drink, smoke, or chew gum for at least 30 minutes before providing a sample.

Saliva collection from donors

For collection by donors at home without medical supervision, kits such as the GeneFix Saliva DNA/RNA range include a simple to attach funnel that makes it easy to collect saliva in a tube prefilled with stabilization reagent for transport and storage of the sample. 


Traditionally, collection devices were designed to collect 1ml or 2ml saliva, but newer devices such as the GFX-03, can collect up to 3ml if higher yields (300μg+) are required, e.g., for biobanking. For maximum yields, donors must ensure that they don’t include bubbles when measuring the liquid level in the tube as this can result in overfilling, or not collecting enough saliva.

Some donors, e.g., children or elderly patients, may find it difficult to produce saliva. If this is the case, saliva production can be stimulated by gently rubbing the cheeks or using a kit such as the SaliFix Saliva Swab DNA Collection kit which contains absorbent swabs for easy collection of smaller volumes of saliva.

Nucleic acid stabilization

One of the key benefits of using saliva as a source of nucleic acids is that samples can be collected remotely and mailed for analysis. Stabilization reagents protect sample integrity by stabilizing DNA and RNA and preventing microbial growth. GeneFix saliva collection kit tubes are prefilled with a non-hazardous guanidium-free stabilization reagent that stabilizes DNA at room temperature for 5 years and RNA for 2 months. Samples can be shipped and stored at room temperature, removing the costs and logistical challenges associated with cold chain storage and transport. For longer term storage GeneFix kits are also suitable for freezing at temperatures down to -80°C.

Safely shipping saliva samples

Shipping packaging and collection tubes must be manufactured from robust materials that can withstand the rigors of the mailing process to ensure your saliva samples arrive at the lab intact. Isohelix sample collection tubes are pressure tested to 95kPa, and Isohelix TPS-50 transport packs are pressure-tested and adhere to UN3373 packaging standards – including secure sealing strips and absorbent material that retains samples leaking from the primary tube within further layers of packaging, preventing sample cross-contamination.

Choosing your DNA/RNA Isolation Kit

Precipitation methods of DNA isolation such as the GeneFix Saliva-Prep Kit, enable the rapid and efficient extraction of high yields of high molecular weight DNA and can be easily scaled to accommodate different sample volumes. As these kits don’t require additional solvents, columns, or filtration, workflows are easy to automate, with fast handling times and reduced sample steps. The unique precipitation chemistry maximizes recovery of high molecular weight DNA, while minimizing co-precipitation of RNA and degraded low-weight DNA.

Alternatively, silica membrane-based spin columns, such as those included in the Xtreme (XME) and Xtreme-RNA (XMR) kits, allow the purification of very high-purity, high molecular weight DNA or RNA for demanding downstream applications such as next generation sequencing or microarrays.

Isolating high molecular weight (HMW) DNA from saliva

Gentle handling is key to preserving the integrity of your DNA samples and isolating HMW DNA for more demanding applications such as whole genome sequencing (WGS) or microarrays. To prevent shearing, use wide bore tips and pipette slowly, avoiding vortexing or shaking tubes.

Cleaning up your DNA after initial extraction

If, despite your best efforts, your extracted DNA is not of a sufficient concentration or purity for your downstream applications, a DNA clean up kit can be used. The Isohelix DNA clean up kit uses a simple 30 minute protocol to reclaim a “failed” sample, preserving your DNA yields and giving High Purity DNA with 260/280 ratios to 1.8.

Nanodrop Scan Before & After DCU CleanUp


By following the advice given in this article, high yields of high purity DNA can be extracted from saliva for use in a number of sensitive downstream applications.

GeneFix offers superior performance and high yields when compared with alternative collectors :

Isohelix Sample Collection: Research highlights from 2023

2023 was a busy year for Isohelix and our customers. In this blog, we present examples of how studies reported in peer-reviewed publications used Isohelix products to solve their sampling challenges.

We’ve highlighted four studies where Isohelix technologies were used to make groundbreaking discoveries in the world of biomedical research. In each of these studies, Isohelix sample collection products were used to collect DNA samples which were then analyzed using highly sensitive genetic analyses such as single nucleotide polymorphism (SNP) genotyping, genome wide association studies (GWAS), and next generation sequencing (NGS) analyses.

  1. GeneFix Saliva Collection Devices are used to collect samples for a large population-based study in Australia
  2. The Hong Kong Genome Project uses GeneFix Saliva Collection Devices to collect saliva samples and the  RapiDri Pouch kit for buccal swabs.
  3. Isohelix saliva collection kits are used to collect saliva from a large cohort of children in the USA
  4. Isohelix Buccal Swabs are used to swab surfaces to collect DNA from bacterial communities in subway facilities

STUDY 1 : Preliminary results from the Australian Genetics of Bipolar Disorder Study: A nation-wide cohort[i]

The Australian Genetics of Bipolar Disorder (BD) Study investigates a nation-wide cohort of adults living with bipolar disorder. The study aims to identify genetic risk factors influencing BD, medication treatment response, and adverse drug reactions (ADRs). Key goals are to characterize the patterns and costs of healthcare usage in BD and to examine the relationship between genetic risk and symptom severity. This article describes the study design and sample characteristics and summarises data collected on BD symptoms, psychiatric comorbidities, and medication use. 

A significant strength of the study is its sample size; it’s the largest population-based study of adults living with BD in Australia. Study participants are based throughout Australia, with many living in rural or remote communities. Saliva samples are collected using Isohelix GeneFixGFX-02 2 mL Saliva Collection Devices which stabilize DNA and RNA at room temperature for up to 60 months,  then mailed to QIMR Berghofer via prepaid Australia Post. Following DNA extraction, genotyping is conducted using the Illumina Global Screening Array V.2.0.


The study demonstrates that Australians with BD are motivated to participate in genetically focused research studies if given the opportunity and provided with simple and effective sample donation protocols. GeneFix Saliva Collectors are ideal for this type of project as their reagents are non-toxic, and the collectors are easy to use as they have a unique funnel design that prevents buffer spillages or flow-back.

The results highlight the high rate of comorbidities and adverse drug reactions among adults living with bipolar disorder in the general Australian population and pave the way for future genomic analyses to identify genetic variants influencing pharmacotherapy treatment response and side effects.

STUDY 2 : The Hong Kong genome project: building genome sequencing capacity and capability for advancing genomic science in Hong Kong[ii]

The Hong Kong Genome Project (HKGP) is the first large-scale genome sequencing (GS) project in the Hong Kong Special Administrative Region. The project aims to sequence 45,000-50,000 genomes in five years, providing participants with diagnoses and personalized treatments and driving the application and integration of genomic medicine into routine clinical care.

For each participant, 6 mL of blood is obtained and stored in two 3-mL EDTA-containing anticoagulation tubes. For buccal swabs, the RapiDri Pouch kit is used to collect cells from inside the cheek, and saliva samples are collected using the GeneFix Saliva DNA Collection and Stabilization Kit. DNA from saliva samples has been shown to be comparable to DNA from blood in many downstream applications[iii] and DNA from saliva samples collected using GeneFix kits is stable at room temperature for over five years, avoiding the high cost and logistical challenges of cold chain transport.

RapiDri Swab

This paper describes the establishment of a robust genome sequencing workflow for the project. The laboratory analyses approximately 350-500 samples per week. During the first 24 months of the project, 12,937 participants and their family members were recruited, and the laboratory has been adapted to facilitate the latest technologies, e.g., long read sequencing and multi-omics.

STUDY 3 : Avoidant Restrictive Food Intake Disorder Genes and Environment (ARFID-GEN): study protocol[iv]

This paper describes the Avoidant Restrictive Food Intake Disorder – Genes and Environment study (ARFID-GEN), which is designed to expand the discovery of genetic and environmental contributions to ARFID risk. The goal of the study is to deliver actionable findings that can be transformed into clinically meaningful insights. This will be achieved by advancing the understanding of the genetic architecture of ARFID relative to other eating disorders and other psychiatric, neurodevelopmental, and metabolic disorders and traits.

3,000 children and adults with ARFID from the United States are included in the study. Efficient online phenotyping of children and adults with ARFID including environmental exposures is combined with at-home saliva sampling, and samples are genotyped to analyze single nucleotide polymorphism (SNP) – based heritability, genome wide association studies, genetic correlations (rgs), and copy number variations (CNVs).


Eligible children are asked to complete a questionnaire on their symptoms and provide a saliva sample using Isohelix saliva collection kits, which are non-toxic and easy for children to use at home as they have a unique funnel design that prevents buffer spillages or flow-back. The kits are provided with return packaging for mailing to the lab for analysis.

The aim is to identify genome-wide significant loci for ARFID, informative rgs, implicated CNVs, and environmental precipitants. Ultimately, the work will hopefully yield information on critical biological pathways that may point toward drug discovery or repurposing that could aid in reversing the tenacity and lethality of these illnesses.

STUDY 4 : Investigation of Surface Bacterial Diversities and Compositions in the Global Subway Facilities[v]

This study explored surface bacterial communities in four global subway facilities using 16S rRNA gene amplicon sequencing of DNA extracted from swabs.

Isohelix Buccal swabs can be used to collect surface DNA from environmental sources as well as from buccal samples. Indoor surface samples for Busan City were collected twice per site using Isohelix DNA/RNA buccal swabs (SK-2S, Isohelix) moistened with sterile 1 mL of phosphate-buffered saline. Samples were collected for 3 min from each site in a 100 cm2 area as recommended in a previous study[vi]. The microbiome was analysed based on the 16S rRNA gene.


Samples collected in Korea were analysed alongside comparable samples obtained from previous studies in Boston[vii], Moscow[viii], and Mexico City[ix].

The study showed that the surface bacterial diversities and taxonomic profiling of the studied subway station surface environments were similar. Most of the predominant phylotypes were Gram-positive microorganisms that probably originated from human and outdoor sources.


The above studies are just a few of the projects that have used Isohelix sample collection and stabilization products to collect samples from research study participants or the environment, for genomic analysis.


Tell us about how Isohelix products helped your study by emailing us at!


[i] Lind, Penelope A., et al. “Preliminary results from the Australian Genetics of Bipolar Disorder Study: A nation-wide cohort.” Australian & New Zealand Journal of Psychiatry 57.11 (2023): 1428-1442.

[ii] Chu, Annie TW, et al. “The Hong Kong genome project: building genome sequencing capacity and capability for advancing genomic science in Hong Kong.”  J Transl Genet Genom 2023;7:196-212

[iii] “New Saliva-Prep2 Isolates High Purity Genomic DNA using GeneFixTM Collectors”, Isoghelix Application Note GSPN: January 2019

[iv] Bulik, Cynthia M., et al. “ARFID Genes and Environment (ARFID-GEN): study protocol.” BMC psychiatry 23.1 (2023): 863.

[v] Kim J, Bae S, Park S, Shukla SK, Yoo K. Investigation of Surface Bacterial Diversities and Compositions in the Global Subway Facilities. Atmosphere. 2023; 14(1):140.

[vi] Jansson, L.; Akel, Y.; Eriksson, R.; Lavander, M.; Hedman, J. Impact of swab material on microbial surface sampling. J. Microbiol. Methods 2020

[vii] Hsu, T.; Joice, R.; Vallarino, J.; Abu-Ali, G.; Hartmann, E.M.; Shafquat, A.; DuLong, C.; Baranowski, C.; Gevers, D.; Green, J.L. Urban transit system microbial communities differ by surface type and interaction with humans and the environment. Msystems 2016, 1, e00018-16

[viii] Klimenko, N.S.; Tyakht, A.V.; Toshchakov, S.V.; Shevchenko, M.A.; Korzhenkov, A.A.; Afshinnekoo, E.; Mason, C.E.; Alexeev, D.G. Co-occurrence patterns of bacteria within microbiome of Moscow subway. Comput. Struct. Biotechnol. J. 2020, 18, 314–322

[ix] Hernández, A.M.; Vargas-Robles, D.; Alcaraz, L.D.; Peimbert, M. Station and train surface microbiomes of Mexico City’s metro (subway/underground). Sci. Rep. 2020, 10, 8798