{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,7,30]],"date-time":"2025-07-30T15:01:28Z","timestamp":1753887688426,"version":"3.41.2"},"reference-count":20,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2021,6,9]],"date-time":"2021-06-09T00:00:00Z","timestamp":1623196800000},"content-version":"vor","delay-in-days":159,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001411","name":"Indian Council of Medical Research","doi-asserted-by":"publisher","award":["5066"],"award-info":[{"award-number":["5066"]}],"id":[{"id":"10.13039\/501100001411","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100007334","name":"Ministry of Health and Family Welfare","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100007334","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["BioMed Research International"],"published-print":{"date-parts":[[2021,1]]},"abstract":"<jats:p>The study is aimed at establishing the optimal parameters for RNA purification of pooled specimens, in SARS\u2010CoV\u20102 assay. This research work evaluates the difference of extracted RNA purity of pooled samples with and without treatment with isopropyl alcohol and its effect on real\u2010time RT\u2010PCR. As per the protocol of the Indian Council of Medical Research (ICMR), 5 sample pools were analysed using qRT\u2010PCR. A total of 100 pooled samples were selected for the study by mixing 50\u2009<jats:italic>\u03bc<\/jats:italic>L of one COVID\u201019 positive nasopharyngeal\/oropharyngeal (NP\/OP) specimen and 50\u2009<jats:italic>\u03bc<\/jats:italic>L each of 4 known negative specimens. Pool RNA was extracted using the column\u2010based method, and 1 set of pooled extracted RNA was tested as such, while RNA of the second set was treated additionally with chilled isopropyl alcohol (modified protocol). Further, the purity of extracted RNA in both the groups was checked using Microvolume Spectrophotometers (Nanodrop) followed by RT\u2010PCR targeting E\u2010gene and RNaseP target. The results showed that the purity index of extracted RNA of untreated pooled specimens was inferior to isopropyl alcohol\u2010treated templates, which was observed to be 85% sensitivity and 100% specificity. The average Cq (E gene) in the unpurified and purified pool RNA group was 34.66 and 31.48, respectively. The nanodrop data suggested that purified RNA concentration was significantly increased with an average value of 24.73 \u00b1 1.49\u2009ng\/uL, which might be the reason for high sensitivity and specificity. Thus, this group testing of SARS\u2010CoV\u20102 cases using pools of 5 individual samples would be the best alternative for saving molecular reagents, personnel time, and can increase the overall testing capacity. However, purity of RNA is one of the important determinants to procure unfailing results, thus, this additional purification step must be included in the protocol after RNA has been extracted using commercially available kit before performing qRT\u2010PCR.<\/jats:p>","DOI":"10.1155\/2021\/6653950","type":"journal-article","created":{"date-parts":[[2021,6,9]],"date-time":"2021-06-09T16:54:40Z","timestamp":1623257680000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Assessment of Successful qRT\u2010PCR of SARS\u2010CoV\u20102 Assay in Pool Screening Using Isopropyl Alcohol Purification Step in RNA Extraction"],"prefix":"10.1155","volume":"2021","author":[{"given":"Mayank","family":"Gangwar","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alka","family":"Shukla","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Virendra Kumar","family":"Patel","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Pradyot","family":"Prakash","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2722-1308","authenticated-orcid":false,"given":"Gopal","family":"Nath","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2021,6,9]]},"reference":[{"key":"e_1_2_10_1_2","unstructured":"https:\/\/covid19.who.int\/."},{"key":"e_1_2_10_2_2","doi-asserted-by":"publisher","DOI":"10.1007\/s13337-020-00599-7"},{"key":"e_1_2_10_3_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0236859"},{"key":"e_1_2_10_4_2","unstructured":"HerperM. BegleyS. BoodmanE. RobbinsR. andFacherL. Thermo Fisher to produce millions of coronavirus diagnostic tests - STAT. 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