Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors
Copyright © 2015 Supachai Sakkhachornphop et al. The 3′-end processing (3′P) of each viral long terminal repeat (LTR) during human immunodeficiency virus type-1 (HIV-1) integration is a vital step in the HIV life cycle. Blocking the 3′P using 3′P inhibitor has recently become an attractive strategy...
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th-cmuir.6653943832-541772018-09-04T10:17:02Z Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors Supachai Sakkhachornphop Weeraya Thongkum Chatchai Tayapiwatana Biochemistry, Genetics and Molecular Biology Immunology and Microbiology Copyright © 2015 Supachai Sakkhachornphop et al. The 3′-end processing (3′P) of each viral long terminal repeat (LTR) during human immunodeficiency virus type-1 (HIV-1) integration is a vital step in the HIV life cycle. Blocking the 3′P using 3′P inhibitor has recently become an attractive strategy for HIV-1 therapeutic intervention. Recently, we have developed a novel real-time PCR based assay for the detection of 3′P activity in vitro. The methodology usually involves biotinylated HIV-1 LTR, HIV-1 integrase (IN), and specific primers and probe. In this novel assay, we designed the HIV-1 LTR substrate based on a sequence with a homology to HIV-1 LTR labeled at its 3′ end with biotin on the sense strand. Two nucleotides at the 3′ end were subsequently removed by IN activity. Only two nucleotides labeled biotin were captured on an avidin-coated tube; therefore, inhibiting the binding of primers and probe results in late signals in the real-time PCR. This novel assay has successfully detected both the 3′P activity of HIV-1 IN and the anti-IN activity by Raltegravir and sodium azide agent. This real-time PCR assay has been shown to be effective and inexpensive for a high-throughput screening of novel IN inhibitors. 2018-09-04T10:09:01Z 2018-09-04T10:09:01Z 2015-01-01 Journal 23146141 23146133 2-s2.0-84929649099 10.1155/2015/853891 https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84929649099&origin=inward http://cmuir.cmu.ac.th/jspui/handle/6653943832/54177 |
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Biochemistry, Genetics and Molecular Biology Immunology and Microbiology Supachai Sakkhachornphop Weeraya Thongkum Chatchai Tayapiwatana Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors |
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Copyright © 2015 Supachai Sakkhachornphop et al. The 3′-end processing (3′P) of each viral long terminal repeat (LTR) during human immunodeficiency virus type-1 (HIV-1) integration is a vital step in the HIV life cycle. Blocking the 3′P using 3′P inhibitor has recently become an attractive strategy for HIV-1 therapeutic intervention. Recently, we have developed a novel real-time PCR based assay for the detection of 3′P activity in vitro. The methodology usually involves biotinylated HIV-1 LTR, HIV-1 integrase (IN), and specific primers and probe. In this novel assay, we designed the HIV-1 LTR substrate based on a sequence with a homology to HIV-1 LTR labeled at its 3′ end with biotin on the sense strand. Two nucleotides at the 3′ end were subsequently removed by IN activity. Only two nucleotides labeled biotin were captured on an avidin-coated tube; therefore, inhibiting the binding of primers and probe results in late signals in the real-time PCR. This novel assay has successfully detected both the 3′P activity of HIV-1 IN and the anti-IN activity by Raltegravir and sodium azide agent. This real-time PCR assay has been shown to be effective and inexpensive for a high-throughput screening of novel IN inhibitors. |
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Supachai Sakkhachornphop Weeraya Thongkum Chatchai Tayapiwatana |
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Supachai Sakkhachornphop Weeraya Thongkum Chatchai Tayapiwatana |
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Supachai Sakkhachornphop |
title |
Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors |
title_short |
Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors |
title_full |
Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors |
title_fullStr |
Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors |
title_full_unstemmed |
Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors |
title_sort |
novel 3′-processing integrase activity assay by real-time pcr for screening and identification of hiv-1 integrase inhibitors |
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2018 |
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https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84929649099&origin=inward http://cmuir.cmu.ac.th/jspui/handle/6653943832/54177 |
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