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"In this book, expert HIV researchers critically review every aspect of this highly evolving and topical subject. The opening chapters deal with the epidemiology, global magnitude and biologic mechanisms of HIV-1 transmission from mother to child through breastfeeding and include considerations of the virus (quantity, compartments, characteristics) and the host (genetic, immunity-innate, cellular, humoral). The effects of breastfeeding on the HIV-infected mother’s health and nutritional status, and the social and cultural issues associated with the practice of breastfeeding are also discussed. The next few chapters provide cutting-edge reviews of the latest approaches to prevention of HIV transmission to the infant through breastfeeding, including antiretroviral strategies, nutritional and immune-based approaches, and treatment of expressed breast milk. The remaining chapters provide a fascinating review of the many iterations this subject has received, as reflected in the several different sets of guidelines for infant feeding by HIV-infected mothers issued by the World Health Organization, and a debate by leading scientists on whether HIV-infected mothers should breastfeed their infants-in resource-limited and in resource-rich settings. A comprehensive overview of the current state of implementing the new evidence for prevention of breastfeeding transmission of HIV all over the world is also presented."

"Serologic assays are commonly used for screening (ELISA) and for confirmation (Western blot) of HIV-1 infection; however, both assays have potentially yielded the false-positive or false-negative results. In this study, a diagnostic RTPCR assay as an alternative test for detection of HIV-1 was developed. Forty-six plasma specimens from highly risky groups, who visited a voluntary counseling and testing for HIV (VCT) in Sanglah Clinic of General Hospital, Denpasar, Bali, were tested by RT-PCR assay with specific primers for Pol region of HIV-1 genome. The results of the RT-PCR tests were then compared with those of serologic tests to obtain the sensitivity and specificity of RT-PCR assay.The results of this study showed that the RT-PCR assay could detect 17 (sensitivity: 65.4%) of 26 serologically positive specimens and was unexpectedly able to detect 2 (specificity: 90%) of 20 serologically negative specimens. Thus, the RT-PCR assay developed in this study is potential to be used as an alternative test, even though there are numerous aspects, particularly the sensitivity, that need to be improved in further research."

"Acquired Immunodeficiency Syndrome disebabkan oleh infeksi Human Immunodeficiency Virus (HIV) terhadap sel-sel T CD4+, limfosit, dan makrofag. Sebanyak 90.7% penderita HIV di Indonesia terinfeksi oleh HIV-1 subtipe CRF01_AE. Protein Gag P7 yang dikode oleh gen gag p7 dan terdapat di dalam genom HIV-1 merupakan protein yang dapat digunakan sebagai obat antiretrovirus dan kandidat antigen untuk sistem diagnostik. Penelitian bertujuan untuk memperoleh fragmen DNA gag p7 (nukleokapsid) HIV-1 yang berukuran 210 pb. Fragmen gen gag p7 diperoleh dari hasil Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) dengan menggunakan primer forward AE_p7F dan primer reverse AE_p7R. Cetakan DNA (template) berasal dari RNA virus yang diambil dari serum pasien penderita HIV-1 di Indonesia. Hasil visualisasi pada gel poliakrilamida 10% menunjukkan bahwa fragmen gen gag p7 (nukleokapsid) berhasil teramplifikasi dengan ukuran 210 pb.

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Acquired immunodeficiency syndrome caused by human immunodeficiency virus infections to the CD4+ cells, lymphocyte, and macrophage. More than 90.7 % HIV patient in Indonesia were infected by HIV-1 subtype CRF01_AE. The Gag P7 protein coded by gag p7 gene which found in HIV genome was known as protein in which could be used for antiretroviral drugs (ARV) and diagnosis of HIV-1 infections. The purpose of research is to get DNA fragment of gag p7 (nucleocapsid) gene of HIV-1 which has length 210 bp. Fragment of gag p7 gene could be gotten from the process of reverse transcriptase polymerase chain reaction (RT-PCR) in which use primer forward AE_p7F and primer reverse AE_p7R. The templates is RNA virus that taken from patient?s serum of HIV-1 in Indonesia. The electrophoresis visualization shown that fragment of gag p7 (nucleocapsid) gene was success amplified in which has length 210 bp."

"Human Immunodeficiency Virus type-I (HIV-1) merupakan penyebab sindroma penurunan sistem imun tubuh yang disebut dengan Acquired Immunodeficiency Syndrome (AIDS). Infeksi HIV-I di dunia dan Indonesia cenderung meningkat. Pemeriksaan yang cepat dan spesifik diperlukan untuk mencegah penyebaran infeksi HIV-I. Berbagai teknik telah dikembangkan untuk deteksi infeksi HIV-I. Pada penelitian ini dikembangkan pemeriksaan RT-PCR HIV-1 Mikrobiologi FKUI (in-house RT-PCR) untuk mendapatkan uji alternatif deteksi HIV-1. Sebanyak 46 plasma dan serum kelompok berperilaku risiko tinggi yang berkunjung ke klinik VCT . RSUP Sanglah Denpasar, telah diperiksa dalam penelitian ini. Serum diperiksa dengan 3 kit rapid test yang berbeda yaitu DetermineTM HIV-1/2 (Abbott), ImmunoCombR HIV 1 & 2 BiSpot (Organics), dan SerodieR HIV-1/2 (Fujirebio Inc.). Plasma diuji dengan pemeriksaan RTPCR generasi I menggunakan primer spesifik terhadap daerah gag dan RT-PCR generasi 2 menggunakan primer spesifik terhadap daerah protease dari genom HIV-1. Hasil rapid test menunjukkan dari 46 sampel, sebanyak 26 serum (56,5%) reaktif dan 20 serum (43,5%) non-reaktif. Tingkat sensitivitas, spesifisitas, nilai duga positif, dan nilai duga negatif RT-PCR generasi 1 secara berturut-turut adalah 80,8%, 95%, 95,5%, dan 79,2%, sedangkan rasio kemungkinan positif dan negatif adalah 16,2, dan 0,2. Pemeriksaan RTPCR generasi 2 menunjukkan tingkat sensitivitas 65,4%, spesifisitas 90%, nilai duga positif 89,5%, nilai duga negatif 66,7%, rasio kemungkinan positif 6,5, dan rasio kemungkinan negatif 0,4. Teknik RT-PCR yang menggunakan primer tersebut dapat mendeteksi HIV pada semua stadium klinis WHO pada kelompok ini. Sensitivitas dan spesifisitas RT-PCR generasi 1 lebih baik daripada RT-PCR generasi 2, tetapi, masih lebih rendah daripada baku emas, Secara keseluruhan, RT-PCR pada penelitian ini belum dapat direkomendasikan sebagai uji altematif baik uji skrining maupun uji konfirmasi dalam mendeteksi infeksi HIV-1.

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Human Immunodeficiency Virus type 1 (HIV-1) can cause decrease of immune response which is called Acquired Immunodeficiency Syndrome (AIDS). HIV-l infection in the world and Indonesia tends to increase. Many techniques were developed to detect HIV-1 infection. A specific and rapid diagnosis is needed to prevent transmission of HIV-1 infection. In this study, we performed RT-PCR HIV-1 Microbiology FKUI (in-house RT-PCR) as an alternative test to detect HIV-1. Forty six plasmas and serums from high risk behavior group who visited VCT Clinic Sanglah General Hospital, Denpasar were used in this study. Serums were tested with 3 different rapid test kits i.e. Determine ° IIIV-112 (Abbott), immunoComb HIV I & 2 BiSpot (Orgenics), and Serodia ' HIV-112 (Fujirebio Inc.). Plasmas were tested with I generation RT-PCR which used specific primers to gag region in HIV-1 genome and specific primers to protease region in IIIV-1 genome for 2nd generation RT-PCR. Results of rapid test demonstrated 26 serums (56.5%) were reactive and 20 serums (43.5%) were non-reactive. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of 1st generation RT-PCR was 80.8%, 95%, 95.5%, 79.2%, whereas positive likelihood ratio (LR +) and negative likelihood ratio (LR -) was 16.2, and 0.2, respectively. The 2"d generation RT-PCR showed sensitivity, specificity, PPV, NPV, LR (+), and LR (-) was 65.4%, 90%, 89.5%, 66.7%, 6.5, and 0.4, respectively. These in-house RT-PCR could detect HIV-1 in all WHO clinical staging in this group. This study showed that lsi generation RT-PCR gives better results than 2"d generation RT-PCR. But still inferior than rapid test to detect HIV-1 infection. Overall, RT-PCR in this study has not been recommended yet as an alternative test to detect HIV-I infection."

"Tujuan Diagnosis cepat dan spesifik seperti uji RT-PCR sangat diperlukan dalam usaha meminimalisasi penyebaran infeksi HIV-1. Oleh karena itu, dalam studi ini dikembangkan uji RT-PCR yang spesifi k terhadap gen gag HIV-1 sebagai target diagnosis. Metode Uji RT-PCR dievaluasi terhadap 46 spesimen yang diperoleh dari voluntary counseling and testing for HIV(VCT) di Rumah Sakit Umum Pmerintah (RSUP) Sanglah, Bali. Untuk mendapatkan sensitivitas dan spesifi tas uji, hasil uji RT-PCR dibandingkan dengan hasil serologi yang umum digunakan di Indonesia. Hasil Uji RT-PCR dapat mendeteksi 21dari 26 spesimen yang positif uji serologi dan memberikan 19 hasil uji negatif dari 20 spesimen yang negatif uji serologi. Satu spesimen menunjukkan hasil positif dengan RT-PCR tetapi negatif dengan uji serologi. Hasil tersebut kemungkinan menggambarkan hasil yang sebenarnya saat uji serologi tidak dapat mendeteksi infeksi HIV-1. Selain itu, lima spesimen yang positif uji serologi menunjukkan hasil negatif dengan RT-PCR yang diduga disebabkan oleh batas deteksi uji RT-PCR yang rendah. Kesimpulan Uji PCR-RT yang dikembangkan dalam studi ini berpotensi digunakan sebagai uji alternatif untuk mendeteksi infeksi HIV-1 dengan 80.0% sensitivitas dan 95.0% spesifisitas.

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AbstractAim A spesifi c and rapid diagnosis such as RT-PCR asay is the most needed to minimize transmission of HIV-1 infection. Therefore, in this study we developed the RT-PCR assay that was spesifi c against the gag gene of HIV-1. Methods The developed RT-PCR assay was evaluated against 46 specimens that were obtained from voluntary counseling and testing for HIV (VCT) in Rumah Sakit Umum Pemerintah (RSUP) Sanglah, Bali. To get the sensitivity and specificity of RT-PCR assay, the results of assays were compared with the results of commercially serologic tests that were commonly used in Indonesia. Results The RT-PCR assay could detect 21 of 26 serologic test-positive specimens and showed 19 negative results of 20 serologic test-negative specimens. There was one specimen that was positive in RT-PCR but negative in serologic assay which might depict a true yield at particular condition when the serologic assay was unable to detect. Five serologic positive-test specimens were negative by RT-PCR that was possibly caused by low detection level of the assay. Conclusion The RT-PCR assay is potential to be used for the detection of HIV-1 infection with a sensitivity and specificity of 80.8% and 95.0% respectively."

"Dalam upaya menginduksi kekebalan berspektrum luas yang responsif terhadap subtipe-subtipe HIV-1 yang berbeda, telah diteliti imunisasi vaksin DNA menggunakan vektor plasmid DNA dan virus fowlpox rekombinan dengan memanfaatkan gen-gen HIV-1 yang dirancang dari runutan konsensus turunan subtipe-subtipe HIV-1 di dunia yang mengekspresikan semua protein dari genom HIV-1 dengan peptida berukuran 30 asam amino yang overlapping dan tersusun secara acak (scrambled antigen vaccines, atau SAVINE).Tiga grup hewan coba yang terdiri dari masing-masing tujuh beruk (Macaca nemestrina) diimunisasi dengan regimen vaksin DNA standar dengan veklor plasmid DNA pHIS-64 dan vektor virus fowlpox rekombinan (rFPV) berbasis gen gag dan pol dan HIV-1 subtipe B, regimen vaksin DNA SAVINE dengan vektor pHIS-64 dan vektor rFPV berbasis genom HIV-1 yang diacak, serta vektor plasmid pHIS-64 dan FPV yang tidak mengandung gen sebagai grup kontrol. Respon kebal selular diamati dengan teknik ELiSpot dan pewamaan silokin intraselular, sedangkan respon kebal humoral diamati dengan teknik ELISA. Pada ketujuh hewan coba yang diimunisasi dengan vaksin DNA HIV-1 standar, secara umum hasil penelitian menunjukkan terinduksinya respon kebal selular terhadap protein Gag HIV-1 serta respon kebal humoral yang ditunjukkan dengan terdeteksinya antibodi terhadap protein p24 HIV-1. Respon kebal selular silang terhadap protein Gag HIV-1 dari subtipe yang berbeda juga ditunjukkan pada grup yang sama. Namun upaya melakukan imunisasi boosting ke-dua dengan vektor rFPV tidak menunjukkan perbaikan induksi respon kebal. Berbeda dari grup hewan coba yang menerima regimen vaksin DNA HIV-1 standar, pada grup yang menerima regimen vaksin DNA HIV-1 SAVINE secara umum tidak menunjukkan adanya induksi respon kebal, kecuali pada satu ekor hewan yang menunjukkan respon kebal selular yang lebih luas terhadap protein Pol dan protein-protein lain HIV-1 meski pada tingkat induksi yang amat rendah. Pengembangan teknologi vaksin SAVINE terus diperbaiki dan disempumakan dengan kemungkinan melibatkan vektor virus aktif yang lain sehingga induksi respon kebal yang diharapkan bisa tercapai.

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T cell immunity plays a critical role in controlling HIV-1 viremia, and encoding a limited set of HIV-1 genes within DNA and poxvirus vectors can, when used sequentially, induce high levels of T cell immunity in primates. However, a limited breadth of T cell immunity exposes the host to potential infection with either genetically diverse HIV-1 strains or T cell escape variants of HIV-1. In an attempt to induce maximally broad immunity, we examined DNA (prime) and recombinant Fowlpox virus (rFPV, boost) vaccines encoding all HIV-1 genes derived from a global HIV-1 consensus sequence, but expressed as multiple overlapping scrambled 30 amino acid segments (scrambled antigen vaccines, or SAVINEs).Three groups of 7 pigtail macaques (Macaca nemestrina) were immunized with sets of DNA and rFPV expressing Gag/Pol antigens only, the whole genome SAVINE antigens, or no HIV-1 antigens. T cell immunity was monitored by ELISpot and intracellular cytokine staining, while the humoral immune response was monitored by p24 antibody capture ELISA. High levels of cross-subtype HIV-specific T cell immunity to Gag were consistently induced in the 7 macaques primed with DNA and rFPV vaccines expressing Gag/Poi as intact proteins. The humoral immunity was also induced in the animals from the same group. It was however, difficult to repeatedly boost immunity with further rFPV immunizations, presumably reflecting high levels of anti-FPV immunity. Unfortunately, this vaccine study did not consistently achieve a broadened level of T cell immunity to multiple HIV genes utilizing the novel whole-virus SAVINE approach, with only one of 7 immunized animals generating broad T cell immunity to multiple HIV-1 proteins. Further refinements are planned with alternate vector strategies to evaluate the potential of the SAVINE technology."

"Dalam upaya menginduksi kekebalan berspektrum luas yang responsif terhadap subtipe-subtipe HIV-1 yang berbeda, telah diteliti imunisasi vaksin DNA menggunakan vektor plasmid DNA dan virus fowlpox rekombinan dengan memanfaatkan gen-gen HIV-1 yang dirancang dari runutan konsensus turunan subtipe-subtipe HIV-1 di dunia yang mengekspresikan semua protein dari genom HIV-1 dengan peptida berukuran 30 asam amino yang overlapping dan tersusun secara acak (scrambled antigen vaccines, atau SAVINE).Tiga grup hewan coba yang terdiri dari masing-masing tujuh beruk (Macaca nemestrina) diimunisasi dengan regimen vaksin DNA standar dengan veklor plasmid DNA pHIS-64 dan vektor virus fowlpox rekombinan (rFPV) berbasis gen gag dan pol dan HIV-1 subtipe B, regimen vaksin DNA SAVINE dengan vektor pHIS-64 dan vektor rFPV berbasis genom HIV-1 yang diacak, serta vektor plasmid pHIS-64 dan FPV yang tidak mengandung gen sebagai grup kontrol. Respon kebal selular diamati dengan teknik ELiSpot dan pewamaan silokin intraselular, sedangkan respon kebal humoral diamati dengan teknik ELISA. Pada ketujuh hewan coba yang diimunisasi dengan vaksin DNA HIV-1 standar, secara umum hasil penelitian menunjukkan terinduksinya respon kebal selular terhadap protein Gag HIV-1 serta respon kebal humoral yang ditunjukkan dengan terdeteksinya antibodi terhadap protein p24 HIV-1. Respon kebal selular silang terhadap protein Gag HIV-1 dari subtipe yang berbeda juga ditunjukkan pada grup yang sama. Namun upaya melakukan imunisasi boosting ke-dua dengan vektor rFPV tidak menunjukkan perbaikan induksi respon kebal. Berbeda dari grup hewan coba yang menerima regimen vaksin DNA HIV-1 standar, pada grup yang menerima regimen vaksin DNA HIV-1 SAVINE secara umum tidak menunjukkan adanya induksi respon kebal, kecuali pada satu ekor hewan yang menunjukkan respon kebal selular yang lebih luas terhadap protein Pol dan protein-protein lain HIV-1 meski pada tingkat induksi yang amat rendah. Pengembangan teknologi vaksin SAVINE terus diperbaiki dan disempumakan dengan kemungkinan melibatkan vektor virus aktif yang lain sehingga induksi respon kebal yang diharapkan bisa tercapai.Specific Immune Responses to the Human Immunodeficiency Virus Type-1 (HIV-1) Proteins In Pigtail Macaque (Macaca nemestrina) Immunized with Whole Gene and Whole Virus Scrambled Antigen Vaccines

T cell immunity plays a critical role in controlling HIV-1 viremia, and encoding a limited set of HIV-1 genes within DNA and poxvirus vectors can, when used sequentially, induce high levels of T cell immunity in primates. However, a limited breadth of T cell immunity exposes the host to potential infection with either genetically diverse HIV-1 strains or T cell escape variants of HIV-1. In an attempt to induce maximally broad immunity, we examined DNA (prime) and recombinant Fowlpox virus (rFPV, boost) vaccines encoding all HIV-1 genes derived from a global HIV-1 consensus sequence, but expressed as multiple overlapping scrambled 30 amino acid segments (scrambled antigen vaccines, or SAVINEs).

Three groups of 7 pigtail macaques (Macaca nemestrina) were immunized with sets of DNA and rFPV expressing Gag/Pol antigens only, the whole genome SAVINE antigens, or no HIV-1 antigens. T cell immunity was monitored by ELISpot and intracellular cytokine staining, while the humoral immune response was monitored by p24 antibody capture ELISA. High levels of cross-subtype HIV-specific T cell immunity to Gag were consistently induced in the 7 macaques primed with DNA and rFPV vaccines expressing Gag/Poi as intact proteins. The humoral immunity was also induced in the animals from the same group. It was however, difficult to repeatedly boost immunity with further rFPV immunizations, presumably reflecting high levels of anti-FPV immunity. Unfortunately, this vaccine study did not consistently achieve a broadened level of T cell immunity to multiple HIV genes utilizing the novel whole-virus SAVINE approach, with only one of 7 immunized animals generating broad T cell immunity to multiple HIV-1 proteins. Further refinements are planned with alternate vector strategies to evaluate the potential of the SAVINE technology."

"Infeksi Human Immunodeficiency Virus tipe I (HIV-1) sebagai penyebab AIDS (Acquired Immunodeficiency Syndrome) merupakan salah satu masalah utama kesehatan dunia yang barns segem diatasi. Sejak ditemukannya penyakit tersebut, vaksin yang dihampkan tidak kanjung tersedia karena berbagai usaha I pengembangan vaksia HIV-1 mengalami hambatan besar oleh karena keanekar&gaman HIV·I yang tinggi. Strategi mutakhir untuk mengatasi hambatan tersebut adalah pengembangan vaksin HIV-I yang spesifik pada subtipe dan populasi di regional tertentu. menggunakan isolat identik dengan sekuen konsensus yang telah ditentukan, sebagai kandidat vaksin.Tujuan pcnelitian ini adalah menentukan sekuen konsensus HlV-1 di Indonesia dengan menggunakan sekuen - sekuen gen protease dan gen reverse transcriptase HN - 1 subtipe paling dominan isola!Indonesia dati isolat damb plasma omng terinfeksi HIV akibat penggunaan narkoba dengan jarum suntik (penasnn). Berdasarkan analisis dalam penelitian ini diketahui bahwa CRFO I_AE merupakan subtipe paling dominan di Indonesia dan telah berhasH diperoleh sekuen konsensus protease dan reverse transcriptase HIV-1 CRFOl_AE Indonesia Sekuen konsensus protease Indonesia tersebut. memiliki perbedaan dengan sekuen konsensus dari database Los Alomos National Laboratory (LANL) sebesar 2,7% untuk sekuen nukleotida (p = 0,030); 5,1% untuk sekuen asam amino (p = 0,000). Sedangkan sekuen konsensus reverse trancriptase Indonesia merniliki perbedaan dengan sekuen konsensus dari LANL sebesar 2,0% nntuk nuklootida (p = 0,208) dan 3,0% untuk asam amiao (p = 0,015).

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Human Immunodeficiency Virus type I (HIV-I) infection as the etiology of AIDS (Acquired Immunodeficiency Syndrome) is a major health problem which need to be urgently solved. Since the discovery of the desease, the effective vaccine is still not available. It is caused by widely the diversity of HIV-I. Novel strategy to overcome this problem is to develop country-specific HIV-1 vaccine, which use the most identical isolate with consensus sequences that had been determined, as vaccine candidate.This study aims to determine consensus sequences (CS) of HIV-1 in Indonesia by using sequences of protease gene and reverse transcriptase l gene of the most predominant subtype HIV-1 sequences from HIV-infected intravenous drog users' blood plasma. This study concluded that CRFOl_AE is I the most predominant subtype HIV-1 in Indonesia Nucleotide and amino acid of Iprotease which determine as CS has 2.7% (p = 0,030) and 5.1% (p = 0,000) differences with CS of CRFOI AE respectively. While nucleotide and amino acid of reverse trancriptase of the CS has 2,0% (p = 0,208) and 3,0".4 (p = 0,015) differences with CS of CRFOI_AE of the LANL, respectively."

"Acquired Immunodeficiency Syndrome (AIDS) merupakan penyakit akibat infeksi Human Immunodeficiency Virus (HIV) yang memiliki jumlah penderita tinggi di Indonesia. Salah satu upaya untuk mencegah bertambahnya jumlah penderita AIDS tersebut ialah dengan penggunaan vaksin. Poliprotein Gag merupakan protein penyusun struktur internal HIV yang dapat digunakan sebagai vaksin karena dapat menginduksi respon imun tubuh. Penelitian telah dilakukan untuk mengekspresikan poliprotein Gag HIV-1 subtipe CRF01_AE yang telah diinsersi ke dalam vektor ekspresi pQE-80L. Ekspresi poliprotein tersebut dilakukan di dalam bakteri Escherichia coli BL21 dan E. coli BL21-CodonPlus (CP) dengan induksi Isoprophyl-β-D-thiogalactopyranoside (IPTG). Pendeteksian poliprotein Gag hasil ekspresi dilakukan dengan metode Sodium Dodecyl Sulphate Polyacrilamide Gel Electrophoresis (SDS-PAGE). Setelah poliprotein berhasil dideteksi, poliprotein Gag kemudian dipurifikasi dengan menggunakan metode kromatografi afinitas Ni2+-NTA di bawah kondisi native. Poliprotein Gag HIV-1 subtipe CRF01_AE dapat diekspresikan dalam E. coli BL21 dan E. coli BL21-CP dengan berat molekul sebesar 55,3 kDa.

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Acquired Immunodeficiency Syndrome (AIDS) is a disease caused by infection of Human Immunodeficiency Virus (HIV) which has a high number of people in Indonesia. One of the efforts to prevent the increasing number of AIDS patients is the use of vaccine. Gag polyprotein is a constituent protein of HIV internal structure that can be used as a vaccine because it can induce immune response of the body. Research has been conducted to express the Gag polyprotein HIV-1 subtype CRF01_AE that have been cloned into the pQE-80L expression vector. Polyprotein expression was carried out in Escherichia coli BL21 and E. coli BL21-CodonPlus (CP) with Isoprophyl-β-Dthiogalactopyranoside (IPTG) induction. Detection of Gag polyprotein was performed by Polyacrilamide Sodium Dodecyl Sulphate Gel Electrophoresis (SDS-PAGE) method. After successfully detected, Gag polyprotein then purified using Ni2+-NTA affinity chromatography under native condition. Gag polyprotein HIV-1 subtype CRF01_AE can be expressed in E. coli BL21 and E. coli BL21-CP with molecular weight is 55,3 kDa."

"Ekspresi gen sintentik gag HIV-1 subtipe CRF01_AE dalam E. coli BL21 dan E. coli BL21-CP telah dilakukan. Gen gag merupakan salah satu gen pada HIV-1 yang tidak mengalami mutasi secara signifikan sehingga gen tersebut dapat digunakan untuk pengembangan vaksin yang dapat dimanfaatkan dalam jangka waktu yang panjang. Pengembangan vaksin HIV membutuhkan protein Gag untuk digunakan sebagai antigen yang mampu merespon pembentukan antibodi pada hewan uji coba. Protein Gag didapatkan dengan cara melakukan ekspresi gen gag yang telah diklon ke dalam vektor ekspresi pQE-81L, dan ditransformasi ke dalam bakteri E. coli BL21 dan E. coli BL21-CP. Ekspresi dilakukan dengan tiga faktor optimasi yaitu, suhu, konsentrasi isopropyl-β-D-thiogalactopyranoside (IPTG) dan waktu ekspresi setelah induksi dilakukan. Analisis hasil ekspresi dilakukan dengan SDS-PAGE dan menunjukkan tidak ada protein Gag yang dihasilkan pada semua keadaan optimasi yang dilakukan. Kegagalan ekspresi gen gag pada E. coli BL21 dan E. coli BL21-CP disebabkan oleh peristiwa kodon bias, dan pemilihan sel inang ekspresi yang kurang tepat.

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Expression of gag gene on HIV-1 subtype CRF01_AE in E. coli BL21 and E. coli BL21-CP had been conducted. Gag gene on HIV-1 is one of the genes that can?t be significantly mutated, so it can be utilized for long term vaccines development. HIV vaccine development requires Gag protein as antigen in order to response antibody formation in animal experiment. Gag protein was obtained by gag gene expression that had been cloned into expression vector pQE-81L and transformed into E. coli BL21 and E. coli BL21-CP. Expression of the gag gene as optimized by temperature, isopropyl-β-D-thiogalactopyranoside (IPTG) concentration, and expression time after IPTG induction. The expression was analyzed by SDS-PAGE and it showed no protein produced in all optimization conditions. The failure of gag gene expression in E. coli BL21 and E. coli BL21-CP caused by codon ray and inappropriate host cell."