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All Journal Jurnal Penelitian Tanaman Industri Narra J Makara Journal of Science
SEKAR ARUMSARI, SEKAR
Mahasiswa Departemen Biokimia, Institut Pertanian Bogor, Kampus IPB Dramaga Gedung Fapet Lantai 5, Jalan Agatis Bogor 16680
Biochemistry, Genetics & Molecular Biology Health Professions Immunology & microbiology Industrial & Manufacturing Engineering Medicine & Pharmacology Public Health

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ANALISIS DIVERSITAS GENETIK AKSESI KELAPA SAWIT KAMERUN BERDASARKAN MARKA SSR TASMA, I MADE; ARUMSARI, SEKAR
853-8212
Publisher : Pusat Penelitian dan Pengembangan Perkebunan

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Abstract

ABSTRAKDiversitas genetik aksesi kelapa sawit Indonesia saat ini sangatrendah. Dalam usaha meningkatkan keragaman genetik telah dilakukaneksplorasi plasma nutfah di pusat keragaman genetik kelapa sawit diKamerun. Tujuan dari penelitian ini untuk mengetahui diversitas genetikdan tingkat polimorfisme berdasarkan marka SSR aksesi-aksesi kelapasawit Kamerun. Bahan tanaman yang digunakan 49 aksesi kelapa sawitKamerun, Afrika yang ditanam di Kebun Sumber Daya Genetik (SDG)Sawit Sijunjung, Sumatera Barat. DNA genomik diisolasi dari tiapindividu aksesi menggunakan protokol isolasi DNA untuk tanamanbergetah. DNA dianalisis menggunakan 20 marka SSR. Dendrogramkekerabatan dikonstruksi menggunakan metode Unweighted Pair GroupMethod Arithmetic (UPGMA) melalui software NTSYS-pc (NumericalTaxonomy and Multivariate Analysis System) versi 2.1-pc. Hasil penelitianmenunjukkan nilai polimorfisme information content (PIC) marka SSRtinggi sebesar 0,80 (berkisar 0,63-0,91). Jumlah alel yang terdeteksi permarka SSR berkisar antara 4-15 alel per lokus SSR (rata-rata 8,75).Analisis filogenetik 49 aksesi menghasilkan diversitas genetik 12,5-54,72% (kemiripan genetik 55,28-87,50%). Pada diversitas genetik54,72%, aksesi Kamerun terbagi menjadi tujuh kelompok masing-masingterdiri dari 9, 28, 4, 2, 1, 2, dan 3 aksesi. Aksesi dengan diversitas genetiktinggi dan berada pada klaster berbeda, potensial digunakan sebagai calontetua dalam program pemuliaan kelapa sawit.Kata kunci: Elaeis guineensis Jacq., diversitas genetik, plasma nutfah,marka SSRABSTRACTGenetic diversity of the Indonesian oil palm collection is very low.To improve their genetic variability, exploration from the oil palm centerof origins has been done in Kamerun. The objectives of this study were todetermine genetic and polymorphism level of the SSR markers Cameroon-originated oil palm accessions. Genetic materials used were 49 Cameroon-originated oil palm accessions collected at Sijunjung Oil Palm GermplamCollection Station, West Sumatera. Genomic DNA was isolated using aprotocol for isolating DNA from leaves rich with latex. DNA was analyzedusing 20 SSR markers. A dendogram was constructed using theUnweighted Pair Group Method Arithmetic (UPGMA) method through theNumerical Taxonomy and Multivariate Analysis System software(NTSYS-pc) version 2.1-pc. Results showed that the polimorfismeinformation content (PIC) values of the SSR markers used was high, 0.80(range from 0.63-0.91). The average number of the SSR alleles detectedwas also high, 8.75 alleles (range from 4-15 alleles per SSR locus).Phylogenetic analysis of the 49 oil palm accessions resulted geneticdiversity of 12.5-54.72% (genetic similarity of 55.28-87.50%). At geneticdiversity 54.72%, the 49 accessions were divided into seven clusters, eachconsisted of 9, 28, 4, 2, 1, 2, and 3 accesions, respectively. Accessionswith high genetic diversity and located at different clusters may be usefulas parent candidates in the future oil palm breeding programs.Key words: Elaeis guineensis Jacq., genetic diversity, germplasm, SSRmarkers
Dual sgRNA-directed knock out survivin gene expression using CRISPR/Cas9 technology for editing survivin gene in triple-negative breast cancer Syahrani, Resda A.; Wanandi, Septelia I.; Arumsari, Sekar; Nihayah, Silviatun; Watanabe, Yukihide; Mizuno, Seiya; Louisa, Melva; Wuyung, Puspita E.
Narra J Vol. 4 No. 3 (2024): December 2024
Publisher : Narra Sains Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.52225/narra.v4i3.1177

Abstract

Clustered regularly interspaced short palindromic repeats (CRISPR)-associated nuclease 9 (CRISPR/Cas9) offers a robust approach for genome manipulation, particularly in cancer therapy. Given its high expression in triple-negative breast cancer (TNBC), targeting survivin with CRISPR/Cas9 holds promise as a therapeutic strategy. The aim of this study was to design specific single guide ribonucleic acid (sgRNA) for CRISPR/Cas9 to permanently knock out the survivin gene, exploring its potential as a therapeutic approach in breast cancer while addressing potential off-target effects. Survivin gene knockout was conducted in the TNBC cell line BT549. Intron 1, exon 2, and intron 2 of the survivin gene were selected as sgRNA targets. These sgRNAs were designed in silico and then cloned into a CRISPR/Cas9 expression plasmid. The cleavage activity was assessed using an enhanced green fluorescent protein (EGFP) expression plasmid. The sgRNAs with higher cleavage activity were selected for the establishment of knockout cells. After transfecting the plasmid into the cells, the success of the survivin gene knockout was validated at the deoxyribonucleic acid (DNA) level using polymerase chain reaction (PCR) and sequencing analysis, and at the protein expression level using Western blotting. The study found that sgRNAs survin1A (targeting intron 1), survex2A (targeting intron 2), and survin2A (targeting intron 2) demonstrated higher cleavage activities compared to the other sgRNAs. However, using the single sgRNA, survex2A did not generate mutations in the survivin gene. At the protein level, survivin was still expressed, indicating that a single sgRNA was ineffective in knocking out the survivin gene. In contrast, the combination of sgRNA survin1A and sgRNA survin2A was more effective in generating mutations in the survivin gene, resulting in the deletion of the entire exon 2 and leading to a loss of survivin protein expression. In conclusion, our work provides specific sgRNAs and demonstrates the utilization of dual sgRNAs strategy in the CRISPR/Cas9 technology to knock out the survivin gene, showing potential in breast cancer therapy.
MnSOD Gene Knockout Promotes Apoptosis in Triple-Negative Breast Cancer BT-549 Cells Through Survivin Inhibition and Caspase-3 and Caspase-9 Modulation Ghanny, Niken Rahmah; Wanandi, Septelia Inawati; Arumsari, Sekar; Paramita, Rafika Indah; Syahrani, Resda Akhra; Putri, Putu Indah Paramita Adi
Makara Journal of Science
Publisher : UI Scholars Hub

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Abstract

Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype characterized by the lack of estro-gen, progesterone, and human epidermal growth factor receptor 2 (HER2) receptors. TNBC cells are becoming more aggressive due to the high expression of manganese superoxide dismutase (MnSOD) antioxidants to suppress reactive oxygen species-induced apoptosis and promote oncogenic signaling. This study was aimed to evaluate the effects of MnSOD knockout (KO) on TNBC cell apoptosis by assessing the survivin, caspase-9, and caspase-3 expressions. BT-549 TNBC cells containing the clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9-edited MnSOD gene were used to evaluate the messenger RNA (mRNA) expressions of survivin using the RT-qPCR assay. The West-ern blot assay was used to measure protein expressions of survivin, caspase-9, and caspase-3. Interactions between MnSOD and apoptosis-related proteins were simulated using computational methods based on molecular docking analysis and protein−protein interaction network. This study revealed that MnSOD KO decreased the binding affinity between MnSOD and survivin, in line with the significant reduction of survivin mRNA and protein expressions whereas the protein expressions of caspase-9 and caspase-3 increased in MnSOD KO cells. Therefore, MnSOD plays a pivotal role in BT-549 cell apoptosis by modulating survivin, caspase-9, and caspase-3 gene expressions. This study provides insights into a novel therapeutic strategy to mitigate the aggressiveness of TNBC by disrupting MnSOD gene expression. Further studies should elaborate the MnSOD signaling pathways involving closely related apoptotic proteins.
Co-Authors Ghanny, Niken Rahmah I Made Tasma Melva Louisa Mizuno, Seiya Nihayah, Silviatun Paramita, Rafika Indah Puspita E. Wuyung Putri, Putu Indah Paramita Adi Septelia I. Wanandi SEPTELIA INAWATI WANANDI Syahrani, Resda A. Syahrani, Resda Akhra Watanabe, Yukihide