DNA Isolation from Mouthwash and Saliva
Mouthwash and salvia are non-invasive ways to collect human cells for genomic analysis. Both mouthwash and saliva contains buccal epithelial cells and white blood cells, which provide rich genetic data. Several companies have their own kits to collect saliva and store it for long periods of time. Another method to collect buccal epithelial cells and white blood cells is mouthwash. As patients are familiar with the use of mouthwash, sampling error is limited compared to similar methods, such as sputum and buccal swabs. Below are two protocols developed by Beckman Life Sciences PoP team members for specific customers and the data when the protocols were used.
DNA Extraction from Saliva Collected in Zeesan Salvia DNA Sample Collection Kit
Genomic DNA (gDNA) was isolated from three healthy donors using Zeesan DNA Salvia DNA Sample Collection Kit. DNA yield was measured by Quant-iT™ PicoGreen™ dsDNA Assay (ThermoFisher). An average of 10.3 μg of gDNA was extracted from 300 μL of saliva [Figure 1]. The error bar represents the standard deviation of three donor replicates.
Purity was accessed on a NanoDrop (Thermo Fisher Scientific). The average A260/280 ratio was 1.9, and the average A260/230 was greater than 2.0 [Figure 2]. The error bar represents the standard deviation of three donor replicates.
DNA integrity was assessed using Agilent Genomic DNA ScreenTape Assay. The DIN scores averaged 7.8 and the peak size of DNA was larger than 55K indicating the gDNA extracted is largely intact [Figure 3 and 4].
DNA Extraction from Mouthwash Samples
Fifteen milliliters of mouthwash containing 12.25% alcohol was given to 5 adults. Due to inherent biological variation three samples were pooled to give technical replicates. The other 2 samples were processed individually. A NanoDrop (Thermo Fisher Scientific) was used to asses yield and purity (Figure 5). The technical replicates gave an average yield of 1.8 µg. The purities for all samples, pooled and individual, gave absorbance ratios sufficient for most downstream applications (Figure 6). The A260/A230 ratios are sufficient given the high amounts of carbohydrates in saliva samples.
Genomic DNA (gDNA) integrity was assessed on an Agilent Genomic DNA Screen Tape (Agilent) (Figure 7). The electropherogram of one of the pooled samples is shown below. The DIN score was 8.1 indicating largely intact DNA.
Beckman Coulter makes no warranties of any kind whatsoever express or implied, with respect to this protocol, including but not limited to warranties of fitness for a particular purpose or merchantability or that the protocol is non-infringing. All warranties are expressly disclaimed. Your use of the method is solely at your own risk, without recourse to Beckman Coulter. Not intended or validated for use in the diagnosis of disease or other conditions. This protocol is for demonstration only, and is not validated by Beckman Coulter.
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