Skrining Marka SSR Untuk Analisis Diversitas Genetik Aksesi Kelapa Sawit - TASMA, I MADE (Buletin Palma, 2014)
Analisis diversitas genetik memfasilitasi pemilihan tetua untuk program pemuliaan kelapa sawit. Penggunaan marka SSR meningkatkan akurasi dan kecepatan analisis filogenetik aksesi kelapa sawit. Sebagai penanda genetik, marka SSR memiliki keunggulan dibandingkan marka DNA lainnya (AFLP, RAPD, dan RFLP) karena marka SSR polimorfisme alelnya tinggi, bersifat kodominan, berdistribusi hampir merata pada genom, ekonomis dalam pengujiannya, dan mampu mendeteksi keragaman genetik aksesi dengan tingkat kekerabatan dekat. Tujuan dari penelitian ini untuk (1) mendapatkan marka SSR yang dapat digunakan untuk analisis diversitas genetik aksesi-aksesi kelapa sawit; (2) menguji marka SSR terpilih untuk uji diversitas genetik dalam aksesi (intra accession), antar aksesi (inter accessions), dan antar species (inter species) kelapa sawit menggunakan marka mikrosatelit terpilih. Sebanyak 39 marka SSR diuji pada empat genotipe kelapa sawit. Pola pita marka SSR diskor dan tingkat polimorfisme marka dihitung. Marka SSR dipilih berdasarkan tingkat polimorfime, pola pita, frekuensi dan ukuran alel terdeteksi, dan penyebarannya pada kromosom kelapa sawit. Sepuluh individu tanaman anggota aksesi C103-T (aksesi Tenera asal Kamerun) dan 9 anggota aksesi E. oleifera (Eo) asal Amerika Selatan dan 7 aksesi E. guineensis (Eg) diuji menggunakan 20 marka SSR terpilih. Dendogram kekerabatan dibuat dengan metode Unweighted Pair Group Method Arithmetic (UPGMA) menggunakan software NTSYS (Numerical Taxonomy and Multivariate Analysis System) versi 2.1-pc. Hasil penelitian menunjukkan 39 marka SSR memiliki tingkat polimorfisme 75% (berkisar 32-92%), jumlah alel 5,97 (2-14) dengan ukuran alel 119-330 bp. Berdasar tingkat polimorfisme, frekuensi dan ukuran alel dan pola pita tersebut, serta distribusinya yang menyebar merata pada kromosom kelapa sawit, terpilih 20 marka SSR untuk diuji selanjutnya. Marka SSR terpilih ini diuji kemanfaatannya untuk analisis keragaman genetik anggota individu dalam aksesi, individu antar aksesi dalam satu species Eo atau Eg, dan antar species Eo dan Eg. Uji filogenetik 20 marka SSR pada 10 individu anggota aksesi C103-T menghasilkan diversitas genetik 14,37-37,38%. Uji 8 individu anggota aksesi Eo menunjukkan diversitas genetik 7-15%. Uji diversitas genetik aksesi antar species Eo dan Eg menghasilkan diversitas genetik sebesar 15-45%. Dengan demikian 20 marka SSR terpilih dapat membedakan dengan sangat jelas anggota individu dalam aksesi, anggota aksesi dalam suatu sepecies, dan anggota aksesi antar species kelapa sawit. Marka SSR terpilih dapat digunakan untuk uji kekerabatan berbagai aksesi kelapa sawit. Hasil studi ini bermanfaat untuk penanganan plasma nutfah di lapang dan pemilihan calon tetua untuk program pemuliaan kelapa sawit. Kata kunci : Kelapa sawit, aksesi, keragaman genetik, marka SSR, uji kekerabatan.ABSTRACTSSR Marker Selection for Genetic Diversity Analysis of Oil Palm AccessionsGenetic diversity analysis of oil palm accessions facilitates parent selection in a breeding program. The use of SSR markers enhances the accuracy and speed of oil palm phylogenetic analysis. SSR is a superior PCR-based genetic marker compared to other developed markers (e.g. AFLP, RAPD, and RFLP) due to its multi allelic nature, codominant characteristic, well-distributed across the plant genome, its economic assays, and high ability in differentiating closely related plant genetic materials. The objectives of this study were to: (1) select the SSR markers appropriate for oil palm accession diversity analysis; (2) analyze the selected markers to be used in genetic relationship studies both in analyzing within- and among- oil palm accessions as well as inter oil palm species. A total of 39 SSR markers were initially used in this study. The appropriate SSR markers were selected based on their polymorphism level, banding pattern, allele frequency and size, together with their distributions across the oil palm genome. The selected markers were used in the phylogenetic analysis studies involving genetic materials within- and among-oil palm accessions. These included 10 individual plants of the C103-T accession (within accession study), 9 accessions of Eo and 7 accessions of Eg (for inter-oil palm accessions and inter species studies). Dendograms were constructed based on Unweighted Pair Group Method Arithmetic (UPGMA) using the Numerical Taxonomy and Multivariate Analysis System (NTSYS) software version 2.1-pc. Results showed that the 39 SSR markers showed polymorphism level of 75% (ranges from 32 to 92%), allele number of 5.97 (2-14), and allele size of 119-330 bp. Based on the observed polymorphism level, allele size and frequency, banding pattern, and their distributions across the oil palm genome, 20 SSR markers were selected to be studied further. The selected SSR markers were tested their robustness to be used in the phylogenetic analyses of individual plants within an accession, among accessions of a species (Eo or Eg) as well as the individuals among oil species Eo and Eg. Phylogenetic analysis of 20 SSR markers on 10 individual plant within a Cameroon originated accession C103-T resulted genetic diversity of 14.37-37.38%. Diversity level of 8 accessions within species Eo was 7-15%. Analysis of inter species diversity Eo and Eg demonstrated diversity level of 5-45%. The observation results indicated that the 20 SSR markers were robust enough to be used in phylogenetic studies of oil palm accessions. The results shown by this study would be very useful in handling the oil palm germplasm in the field and should facilitate parent selection in a breeding program