| dc.description.abstract | Major objective of the study: To develop an in-house RNA extraction kit for extraction of high quality RNA.
Specific objectives of the study: To determine the reagents required to develop the in-house RNA
Extraction kit, to develop an inhouse RNA extraction kit for the extraction of high-quality RNA,
and to compare the performance of the developed kit with a commercial extraction kit.
Methodology: The study was at the Genomics and Molecular biology laboratory, College of
Health Sciences Makerere University between April 2023, and June 2023. It was an applied
experimental study aimed at developing and evaluating the extraction efficiency of an in-house
SARS-CoV-2 RNA extraction kit. Reagents and samples used for the study were obtained from
the Genomics and Molecular biology laboratory with permission obtained from the laboratory
management. Nine nasopharyngeal swab samples for covid-19 that were stored under
appropriate conditions were used for the study being randomly sampled to contain both positive
and negative samples. SARS-CoV-2 RNA extraction from the 9 samples was done in two sets
using two RNA extraction kits i.e., developed in-house covid-19 RNA extraction kit and the
Zymo Quick Viral RNA kit which acted as the reference kit for comparison during extraction
efficiency assessment. A PCR master mix was prepared using the available commercial RT-PCR
kit [Novel Coronavirus (2019-nCoV) Real Time Multiplex RT-PCR] kit, and test run was
performed using Quant Studio 7 Flex PCR-platform.
Results: In this study, viral RNA was extracted from nine samples using two different kits, an in house developed kit, and the Zymo Quick Viral RNA kit. Out of the Nine samples extracted with
the inhouse developed kit, seven samples tested negative for viral RNA and two tested positive.
Meanwhile the same samples processed with the Zymo Quick Viral RNA kit, five samples tested
negative, and four samples tested positive for SARS Cov-2 viral RNA. There were discrepancies
in the results obtained that could have been due to extraction errors, contamination and even
errors during preparation for the PCR. Nevertheless, the general ability of the developed kit to
extract covid-19 viral RNA was evident. | en_US |
