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All Journal VALENSI International Journal of Renewable Energy Development Jurnal Riset Kimia Jurnal Keramik dan Gelas Indonesia Science and Technology Indonesia Al-Kimia Chimica et Natura Acta ALCHEMY Jurnal Penelitian Kimia Indonesian Journal of Chemistry PENDIPA Journal of Science Education Molekul: Jurnal Ilmiah Kimia Jurnal Keramik dan Gelas Indonesia Journal of Advanced Technology and Multidiscipline (JATM)
Yeni Wahyuni Hartati
Departemen Kimia, Fakultas Matematika Dan Ilmu Pengetahuan Alam, Universitas Padjadjaran
Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Chemical Engineering, Chemistry & Bioengineering Chemistry Civil Engineering, Building, Construction & Architecture Computer Science & IT Education Electrical & Electronics Engineering Energy Environmental Science Industrial & Manufacturing Engineering Materials Science & Nanotechnology Mathematics Physics

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In-house Multiplex rRT-PCR Assay for Sars-Cov2 Detection in Indonesia using a new primer design Gaffar, Shabarni; Putri, Rafika Nanda; Shabrinna, Hanif; Supriyadi, Isma Yustifania; Wiraswati, Hesti Lina; Ekawardani, Savira; Ishmayana, Safri; Hartati, Yeni Wahyuni; Faridah, Lia
Jurnal Riset Kimia Vol. 16 No. 1 (2025): March
Publisher : Universitas Andalas

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.25077/jrk.v16i1.781

Abstract

During the COVID-19 pandemic, we attempted to develop an in-house rRT-PCR kit, utilizing custom primers targeting NSP14 and RdRp, with the RPP30 gene as an internal control. This kit will support Indonesian independence in enhancing COVID-19 diagnostics. The primer and probe were designed by a bioinformatics tool, determining primer specificity and sensitivity, optimizing probe concentrations, establishing LoD (Limit of Detection), LoQ (Limit of Quantification), and rRT-PCR efficiency, multiplex testing of the rRT-PCR kit on clinical samples, and testing the kit's stability. The in-house rRT-PCR kit can detect NSP14, RdRp, and RPP30 genes. The optimal concentrations for the NSP14, RdRp, and RPP30 probes are 1 μM, 1.5 μM, and 1.5 μM, respectively. The LoD and LoQ for the NSP14 are 0.22 ng/μL and 0.67 ng/μL, and for the RdRp are 1.08 ng/μL and 3.28 ng/μL. The rRT-PCR efficiencies for the NSP14, RdRp, and RPP30 are 80.3%, 100.6%, and 106%, respectively. Detection of ten clinical samples, comprising seven true positive and three true negative samples, showed Ct (Cycle threshold) values of 28–31 for the RPP30 gene, Ct 21–27 for the RdRp gene, and Ct 30–34 for the NSP14 gene. Stability testing of the rRT-PCR kit demonstrated promising results, where the kit stored at -20°C for seven days showed almost no difference in Ct values. This in-house multiplex rRT-PCR will support Indonesian independence in enhancing COVID-19 diagnostics, providing a dependable method for detecting SARS-CoV-2 in clinical samples.
Co-Authors Afrizal Mayub Aga Adi Masyhuri Amelia Shafira Annisa Ilma Naviardianti Annisa Ilma Naviardianti Annisa Luthfiah Ari Hardianto Arum Kurnia Sari Atiek Rostika Noviyanti Azhari Yusuf Azizah, Mamlikatu Ilmi Budi Adiperdana Dewi Handayani Diana Hendrati Diana Hendrati Diana R. Eddy Diana Rakhmawaty Eddy Diding Mandala Putra Egista Istioka Fazrin Ekawardani, Savira Elin Marlin Elsa Nur Utami Fadhilah, Riana Adisti Firdaus, Muhammad Lutfi Ghina Nur Fadhilah Hesti Lina Wiraswati Husein Hernadi Bahti Husein Hernandi Bahti Iman Permana Maksum Irkham Irkham Irkham, Irkham Khaerani, Wulan Lena Rahmidar Lia Faridah Lubis, Rubianto A. M. Lutfi Firdaus M. Lutfi Firdaus M. Lutfi Firdaus Mahendra Permadi Masyhuri, Aga Adi Muhamad Diki Permana Muhammad Hilman Daniswara Muhammad Yusuf Mulyani, Rahmaniar Nida Yumna Nurmalasari, Ratna Nursaadah, Euis Nurul Auliany Pratomo, Uji PUTRI, RAFIKA Putri, Rafika Nanda Rahmaniar Mulyani Ravenna Aristantia Riana Adisti Fadhilah Rini Surbakti Rizki M. Okumura Rizki Muharami Cedia Sari Rubianto A. Lubis Rukiah Rukiah Rustaman Rustaman Safri Ishmayana Salma Nur Zakiyyah Santhy Wyantuti Sari Syahruni Savira Ekawardhani Shabarni Gaffar Shabrinna, Hanif Shauvina, Shauvina A Sheila Destiani Solihudin Solihudin Supriyadi, Isma Yustifania Titin Sofyatin Toto Subroto Uji Pratomo Yohan Yohan Yohan Yohan Zakiyyah, Salma Nur Zalfadilah, Faizal Nur Zein, Muhammad Ihda Hamlu Liwaissunati