High-yield RNA-extraction method for saliva

Pandit, Pratibala, Cooper-White, Justin and Punyadeera, Chamindie (2013) High-yield RNA-extraction method for saliva. Clinical Chemistry, 59 7: 1118-1122. doi:10.1373/clinchem.2012.197863

Author Pandit, Pratibala
Cooper-White, Justin
Punyadeera, Chamindie
Title High-yield RNA-extraction method for saliva
Journal name Clinical Chemistry   Check publisher's open access policy
ISSN 0009-9147
Publication date 2013-07
Year available 2013
Sub-type Article (original research)
DOI 10.1373/clinchem.2012.197863
Volume 59
Issue 7
Start page 1118
End page 1122
Total pages 5
Place of publication Washington, DC United States
Publisher American Association for Clinical Chemistry
Collection year 2014
Language eng
Formatted abstract
BACKGROUND: The use of salivary diagnostics is increasing because of its noninvasiveness, ease of sampling, and the relatively low risk of contracting infectious organisms. Saliva has been used as a biological fluid to identify and validateRNAtargets in head and neck cancer patients. The goal of this study was to develop a robust, easy, and cost-effective method for isolating high yields of total RNA from saliva for downstream expression studies.

Oral whole saliva (200 μL) was collected from healthy controls (n = 6) and from patients with head and neck cancer (n = 8). The method developed in-house used QIAzol lysis reagent (Qiagen) to extract RNA from saliva (both cell-free supernatants and cell pellets), followed by isopropyl alcohol precipitation, cDNA synthesis, and real-time PCR analyses for the genes encoding β-actin ("housekeeping" gene) and histatin (a salivary gland-specific gene).

RESULTS: The in-house QIAzol lysis reagent produced a high yield of total RNA (0.89 -7.1 μg) from saliva (cell-free saliva and cell pellet) after DNase treatment. The ratio of the absorbance measured at 260nmto that at 280 nmranged from 1.6 to 1.9. The commercial kit produced a 10-fold lower RNA yield. Using our method with the QIAzol lysis reagent, we were also able to isolate RNA fromarchived saliva samples that had been stored without RNase inhibitors at -80°C for >2 years.

Our in-house QIAzol method is robust, is simple, provides RNA at high yields, and can be implemented to allow saliva transcriptomic studies to be translated into a clinical setting.
Keyword Transcriptome
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

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