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Analisis Keragaman Genetik Tetua Dan Hasil perpustakaan.uns.ac.id digilib.uns.ac.id ANALISIS KERAGAMAN GENETIK TETUA DAN HASIL PERSILANGAN ANGGREK HITAM (Coelogyne pandurata Lindl.) DISERTASI Disusun Untuk Memenuhi Sebagian Persyaratan Mencapai Gelar Doktor Program Doktor Ilmu Pertanian Oleh SRI HARTATI NIM T. 651108006 PROGRAM DOKTOR ILMU PERTANIAN PROGRAM PASCASARJANA UNIVERSITAS SEBELAS MARET SURAKARTA commit to201 user5 i perpustakaan.uns.ac.id digilib.uns.ac.id ANALISIS KERAGAMAN GENETIK TETUA DAN HASIL PERSILANGAN ANGGREK HITAM (Coelogyne pandurata Lindl) DISERTASI Oleh SRI HARTATI NIM T 651108006 Tim Penguji Jabatan Nama Tanda Tangan Ketua Prof. Dr. Ravik Karsidi, M.S. NIP 195707071981031006 ……………… Sekretaris Prof. Dr. Ir. Ahmad Yunus, MS NIP 196107171986011001 ……………. Anggota Prof. Dr. Ir. Bambang Pujiasmanto, MS Penguji NIP 195602251986011001 …………… Dr. Ir. Supriyadi, M.S. NIP 195813081985031003 …………… Prof. Dr. Ir. Nandariyah, M.S. NIP 195408051981032002 …………… Prof. Dr.Ir. Ahmad Yunus, M.S. NIP 196107171986011001 …………… Dr. Ir. Djati Waluyo Djoar, M.S. NIP 195102021980031003 …………… Dr. Ir. Parjanto, M.P. NIP 196203231988031001 …………… Dr. Ir. Widyatmani Sih Dewi, M.P. NIP 196311231987032002 …………… Prof. Dr. Ir. Y. Purwanto, DEA NIP 196102181985031003 Telah dipertahankan di depan penguji pada sidang Ujian Disertasi dan dinyatakan telah memenuhi syarat pada tanggal 26 Maret 2015 Mengetahui Rektor Universitas Sebelas Maret Surakarta Prof.commit Dr. toRavik user Karsidi , M.S. NIP 195707071981031006 ii perpustakaan.uns.ac.id digilib.uns.ac.id PERNYATAAN KEASLIAN DISERTASI DAN PUBLIKASI DISERTASI Saya menyatakan sebenar-benarnya bahwa : Disertasi yang berjudul “Analisis Keragaman Genetik Tetua Dan Hasil Persilangan Anggrek Hitam (Coelogyne pandurata Lindl.)” ini adalah karya ilmiah saya sendiri dan tidak terdapat isi karangan yang pernah diajukan oleh orang lain untuk memperoleh gelar akademik serta tidak terdapat karya atau pendapat yang diterbitkan oleh orang lain, kecuali yang digunakan sebagai acuan yang disebutkan sumbernya, baik dalam naskah karangan dan daftar pustaka. Apabila ternyata dalam naskah disertasi ini dapat dibuktikan terdapat unsur-unsur plagiasi, maka saya bersedia menerima sangsi dan diproses sesuai dengan peraturan perundangundangan yang berlaku (UU No. 20 Tahun 2003 pasal 25 ayat 2 dan pasal 70). Publikasi sebagian atau keseluruhan isi disertasi pada Journal atau frum ilmiah harus menyertakan tim promotor sebagai author dan PPs UNS sebagai institusinya. Apabila saya melakukan pelanggaran dari ketentuan publikasi ini, maka saya bersedia mendapatkan sangsi akademik yang berlaku. Surakarta, Maret 2015 Mahasiswa, Sri Hartati NIM T. 651108006 commit to user iii perpustakaan.uns.ac.id digilib.uns.ac.id KATA PENGANTAR Puji dan syukur senantiasa penulis panjatkan kepada Allah SWT atas Rahmat dan HidayahNya sehingga penulis dapat menyelesaikan penelitian dan penulisan disertasi yang berjudul “ANALISIS KERAGAMAN GENETIK TETUA DAN HASIL PERSILANGAN ANGGREK HITAM (Coelogyne pandurata Lindl.)”. Sejak dimulainya penelitian, hingga selesai penulisan disertasi ini penulis banyak mendapatkan bantuan baik berupa bimbingan, bantuan moral material, gagasan yang kesemuanya sangat bermanfaat bagi penulis. Terima kasih yang mendalam penulis ucapkan kepada yang terhormat: 1. Prof. Dr. Ravik Karsidi, MS. selaku Rektor Universitas Sebelas Maret sekaligus sebagai ketua tim penguji ujian terbuka. 2. Prof. Dr. Ir. Ahmad Yunus, MS, Direktur Pascasarjana merangkap ko- promotor yang telah banyak memberikan bimbingan dan saran penelitian sampai penyelesaian disertasi ini. 3. Prof. Dr. Ir. Bambang Puji Asmanto, MS, Dekan Fakultas Pertanian yang telah memberikan ijin belajar dan tim penguji. 4. Prof. Dr. Ir. Nandariyah, MS, selaku Promotor yang dengan penuh kearifan telah memberikan bimbingan, nasehat, petunjuk arahan sejak dari perencanaan dan selama pelaksanaan penelitian sampai penyelesaian disertasi ini, 5. Dr. Ir. Djati Waluyo Djoar, MS, selaku ko-promotor yang dengan penuh kesabaran, pengertian dan ketulusan telah banyak memberikan bimbingan dan, dorongan moral yang sangat berguna dalam pelaksanaan penelitian sampai selesainya penulisan disertasi ini, 6. Dr. Ir. Supriyadi, MS selaku Ketua Program Studi S3 Ilmu Pertanian dan tim penguji. 7. Dr. Ir. Widyatmani Sih Dewi, M.P. selaku sekertaris Program Studi S3 Ilmu Pertanian dan tim commitpenguji .to user iv perpustakaan.uns.ac.id digilib.uns.ac.id 8. Prof (Riset) Dr. Ir. Y. Purwanto DEA, dari Pusat Penelitian Biologi LIPI Bogor selaku penguji yang banyak memberikan masukan berharga. 9. Dr. Ir. Parjanto MP selaku tim penilai dan penguji 10. Ir. Susilo Hambeg Poromarto MSi., PhD. selaku tim penilai 11. Prof. Dr. Ir. Edi Purwanto MSc selaku penilai seminar hasil. 12. Kepala Pusat Konservasi Tumbuhan Kebon Raya LIPI Bogor, 13. Koordinator Kebon Anggrek Kebon Raya LIPI Bogor Ir. Dwi Murti Puspitaningtyas, MSi dan Dr. Ir. Joko Ridho atas fasilitas kebun dan materi yang disediakan. Sdr. Ponco Yulianto, Bu Yuniar, Pak Supardi, Bu Yupi, Bu Liza, Bu Sutini yang telah banyak membantu pelaksanaan di lapang. 14. Kepala Laboratorium Genetika Pusat Penelitian Biologi LIPI Bogor Dr. Ir. Yuyu S. Poerba, Teknisi Lab Genetika LIPI Bogor: Bu Tri, Mbak Herlina, Bapak Fajar, dan Pak Hafid (Lab Sitologi –LIPI Bogor). 15. Prof. Sobir dan mbak Sulasih dari Pusat Kajian Hortikultura dan Tropika IPB Bogor atas ijin pemakaian Laboratorium Molekuler. 16. Teman-teman kuliah S3 Ilmu Pertanian angkatan 2011: Ir. Sukaya, MS., Ir. Edi Tri Haryanto, MP., Ir. Endang Setyo Mulyo, MSi., Ir. Dwi Harjoko, MP., Dra. Farida Yuliani, MS., Ir. Priyono, MP., Ir. Catur Sulistyorini, MM. yang telah memberi dukungan dan semangat untuk menyelesaikan disertasi ini. 17. Sdr. Thithin, Riya, Linda, Hespriawan mahasiswa Agroteknologi Fakultas Pertanian UNSyang telah ikut membantu pelaksanaan penelitian. 18. Penghormatan dan ucapan terima kasih yang khusus disampaikan kepada suami tercinta Prof. Dr. Ir. Ongko Cahyono, M.Sc dan semua anakku Doan Perdana ST, MT dan istrinya Imas Nurliah, dr. Febrian Dwi Cahyo Sp.An. M. Kes. dan istrinya dr. Hanindia Riani Prabaningtyas serta Adiptya Cahya Mahendra S. Ked. Atas semua pengorbanan dukungan moril dan matertiil serta kasih sayang yang tulus selama kuliah S3 Ilmu Pertanian di Pasca Sarjana UNS. Penulis menyadaricommit bahwa to disertasi user ini masih jauh dari sempurna, v perpustakaan.uns.ac.id digilib.uns.ac.id karena kesempurnaan itu hanya milik Allah SWT sehingga saran dan kritik yang bersifat membangun dalam menyempurnakan disertasi ini sangat penulis harapkan. Akhirnya penulis berharap disertasi ini dapat dapat bermanfat untuk perkembangan anggrek di Indonesia. (Sri Hartati) commit to user vi perpustakaan.uns.ac.id digilib.uns.ac.id ABSTRAK Anggrek hitam (Coelogyne pandurata Lindl.), merupakan salah satu anggrek langka Kalimantan Timur yang memiliki kekhasan bunga besar, berwarna hijau dengan lidah berwarna hitam. Anggrek ini perlu dilestarikan melalui persilangan dengan spesies lain. Penelitian ini bertujuan untuk (1) mendapatkan informasi keragaman genetik anggrek Coelogyne spp dan menentukan kedekatan genetik antara anggrek hitam dengan spesies lain dalam genus Coelogyne berdasarkan karakter morfologi dan molekuler RAPD (Random Amplified Polymorphic DN. (2) mendapatkan metode persilangan yang kompatibel antara C. pandurata dengan tetua terpilih. (3) mendapatkan informasi karakter sitologi (kromosom) dan tingkat ploidi pada F1 hasil persilangan antara C. pandurata dengan tetua terpilih. (4) mendapatkan informasi besarnya keragaman baru pada F1 hasil persilangan C. pandurata berdasarkan penanda molekuler RAPD dan ISSR (InterSimple Sequence Repeats). Bahan yang digunakan adalah tanaman anggrek koleksi Pusat Konservasi Tumbuhan Kebon Raya LIPI Bogor yaitu Coelogyne spp. meliputi: C. pandurata, C. massangeana, C. mayeriana, C. asperata, C. celebensis dan C. rumphii. Penelitian dilakukan dalam lima kajian yaitu: (1) Identifikasi keragaman genetik anggrek Coelogyne spp berdasarkan penanda morfologi. Identifikasi dilakukan secara deskriptif dengan menggunakan 45 karakter; (2) Identifikasi keragaman genetik anggrek Coelogyne spp berdasarkan penanda molekuler RAPD. Identifikasi dilakukan dengan menggunakan 11 macam primer; (3) Teknik hibridisasi untuk menambah ragam genetic anggrek hitam meliputi tiga metode: crossing (♀ C. pandurata x ♂ C. rumphii), reciprocal (♀ C. rumphii x ♂ C. pandurata) dan selfing yaitu polinia ditransfer ke dalam stigma pada satu bunga dalam satu tanaman; (4) Identifikasi hasil persilangan anggrek C. pandurata. dengan C. rumphii berdasarkan analisis sitologi menggunakan metode Squasing dan flow cytometry menggunakan alat Partec CyFlow space (Partec GmbH); (5) Identifikasi hasil persilangan anggrek C. pandurata dengan C. rumphii berdasarkan molekuler RAPD dan ISSR menggunakan 6 primer RAPD dan 4 primer ISSR Analisis data dilakukan sebagai berikut. skoring data morfologi dilakukan dari hasil deskripsi menjadi data biner. Data molekuler diamati dengan menentukan skor berdasarkan ada atau tidaknya pita DNA. Pita DNA diterjemahkan dalam data biner, jika ada nilai 1 dan jika tidak ada nilai 0. Analisis klaster/gerombol dilakukan dengan program NTSYSpc versi 2.02i dengan metode UPGMA (Unweighted Pair Group Method of Arithmatic Average) fungsi SimQual (Rohlf , 2000). Hasil penentuan keragaman baik secara morfologi maupun secara molekuler dari enam spesies anggrek dari genus Coelogyne spp diperoleh commit to user karakter yang beragam secara morfologi antara 2% – 22% dan secara vii perpustakaan.uns.ac.id digilib.uns.ac.id
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