Judul: Isolasi Konstruksi dan Rekombinasi Gen Pengkode Reverse Transcriptase asal Simian Retrovirus-2 (SRV-2)
Abstrak: The availability of reverse transcriptase enzym as a valuable component for Reverse Transcription Polymerase Chain Reaction (RT-PCR) technique has been providing by foreign companies. Therefore, it needs more cost and time to provide it. This study is an early step to produce in house reverse transcriptase rekombinant enzym from SRV-2. The purpose of this study was isolated, constructed, and rekombined SRV-2 reverse transcriptase gene to entry clone and destination vector. Isolation was done by PCR amplification technique using three pairs of primers, which are RT I (3282-3683), RTII (3261-3900), and RTIII (3169-4133). Construction and recombination in this study used Gateway method from Invitrogen. SRV-2 reverse transcriptase gene was isolated in this study with A-549 cell inoculated SRV-2 as source of virus. This gene amplify by PCR technique and resulted specific band for RTI 401 bp, RTII 639 bp, and RTIII 964 bp. Analysis of pENTR/SD/TOPO and pDEST17 indicated SRV-2 reverse transcriptase gene code has already inserted to this plasmid. This result indicated SRV-2 reverse transcriptase gene have isolated, constructed, and recombined to entry clone and destination vector.
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Judul: Pengembangan enzim rekombinan reverse transcriptase simian betaretrovirus serotipe-2 isolat indonesia asal monyet ekor panjang
Abstrak: Reverse transcriptase (RT) is a multifunctional enzyme that catalyzes the formation of double-stranded DNA from single-stranded RNA viral genomes. Reverse transcription is an essential and critical step in life cycle of all retroviruses replication. RT has become an important enzyme in molecular biology, genetics and medicine for the synthesis of complementary DNA (cDNA) from messenger RNA (mRNA). The application of this enzyme combining with PCR amplification technique (RT-PCR) has expanded our knowledge of numerous cellular regulatory mechanisms and has become a gold standard to facilitate the coding region cloning of any gene of interest. The development of biotechnology has been relying on the availability of this efficient RT. In biomedical fields, RT of human immunodeficiency virus (HIV), the virus causing acquired immunodeficiency syndrome (AIDS) has been the most studied enzyme due to the prime target for the development of antiretroviral drug therapy of HIV/AIDS. Simian betaretrovirus serotype-2 (SRV-2) is the causative agent of simian acquired immunodeficiency syndrome (SAIDS) in Asian macaques with varying severity. While SRV is an important pathogen in Asian macaque and causes a potential confounding variable in biomedical research, SRV also provides a valuable viral model to compare with other retrovirals for a better understanding of aspects of retroviral–host interactions, infection mechanism, retroviral structure, antiretroviral and vaccine development. As is characteristic of all retroviruses, SRV-2 can transcribe its own RNA genome into a double-stranded DNA by the action of reverse transcriptase enzyme. The presence of gene encoding RT enzyme in SRV-2 genome potentially be studied and utilized further as RT enzyme model for developing the anti RT drug of others retrovirus. On the other hands, the gene encoding SRV-2 RT can be isolated and cloned to produce RT recombinant enzyme and applied in molecular biology techniques. In this study, we isolated the gene encoding RT enzyme of SRV-2 that infected Indonesian cynomolgus monkey and predicted the three dimensional (3D) structure model using I-TASSER (Iterative Threading ASSEmbly Refinement) computational program and visualized with PyMol program. Gene expression was constructed in Escherichia coli expression system with Gateway technology methods using pENTR/SD/TOPO entry vector and pDEST17 destination vector. Purification was performed using Ni2+-NTA affinity chromatography. Expression was analyzed using SDS-PAGE and Western blot techniques; meanwhile enzyme activity was measured using RT colorimetric assay and RT-PCR technique. We isolated the gene encoding RT enzyme of SRV-2 Indonesian isolate at nucleotide position 3284-4925 consisted of 547 amino acids. The I-TASSER program has predicted the three dimensional structure model of RT SRV-2 enzyme that consisted of palm/finger, thumb, connection and RNase H domain. The polymerase active site located in the finger/palm subdomain characterized by three conserved catalytic aspartates (Asp90, Asp165, Asp166), and has a highly conserved YMDD motif as Tyr163, Met164, Asp165, and Asp166. This SRV-2 RT structure model has the highest homology to HIV-1 RT (2B6A.pdb). Meanwhile, the structure of SRV-2 RNase H closely related to its ortologs such as Escherichia coli RNase H (1GOA) and HIV-1 RNase H (3QIO, 3HYF). The catalytic active site of SRV-2 RNase H contains three amino acid carboxylateresidues as Asp436, Glu465 and Asp486. Analysis of expression using SDS
Keyword: reverse transcriptase, recombinant enzyme, SRV-2 RT,, Escherichia coli expression system, I-TASSER

Judul: Keawetan Alami Kayu Mangium (Acacia mangium Willd.) Umur 5, 6, dan 7 Tahun
Abstrak: Penelitian ini bertujuan untuk melakukan pengujian keawetan alami kayu berupa uji kubur kayu mangium pada umur 5, 6, dan 7 tahun serta berbagai posisi kayu. Metode yang diterapkan adalah uji kubur mengacu kepada ASTM D 1758- 06. Hasil penelitian menunjukkan kehilangan berat terhadap umur kayu tertinggi pada umur 5 tahun sebesar 34.13% dan terendah pada umur 7 tahun sebesar 14.02%. Kehilangan berat terhadap posisi kayu terendah terdapat pada bagian pangkal kayu sebesar 11.16%, sedangkan kehilangan berat tertinggi terdapat pada bagian ujung kayu sebesar 29.00%. Analisis statistik menyimpulkan bahwa umur kayu, posisi kayu di pohon, serta interaksi antara umur dan posisi kayu di pohon terhadap kehilangan berat berbeda nyata. Umur 7 tahun posisi pangkal memiliki kehilangan berat yang paling kecil, sedangkan umur 5 tahun posisi ujung memiliki kehilangan berat yang paling besar. Kelas mutu kayu mangium yang mengacu kepada ASTM D1758-06 pada umur 5 tahun yaitu kelas mutu 4, kelas mutu 7 pada umur 6 tahun, dan kelas mutu 6 pada umur 7 tahun. Posisi kayu pangkal dan tengah memiliki kelas mutu 6 sedangkan posisi kayu ujung memiliki kelas mutu 4. Pengujian kondisi lingkungan menunjukkan suhu lingkungan, suhu dalam tanah, serta kelembapan sebesar 28.10 oC, 25.10 oC, dan 78.10 %.
Keyword: keawetan alami, mangium, posisi kayu, umur