SummaryIn temporary immersion system (TIS),plant materials are exposed to the medium fora short time, therefore they are more exposedto the air and a lack of oxygen frequentlyexperienced by a liquid culture can be avoided.This experiment was conducted to determinethe procedure for callus proliferation up tosomatic embryo germination of oil palm(Elaeis guineensis Jacq.) in TIS culture.Embryogenic calli of oil palm clone MK 638from Marihat Research Institute were culturedon solid medium in the dark culture room andthen used as materials for TIS. Immersion timefor all cultures was three minutes every sixhours. Callus proliferation was conducted inDF liquid culture with 5 mg/L 2,4-D and0.1 mg/L kinetin with transfer interval of 4, 6and 8 weeks. The treatments for somaticembryo maturation were kinetin and ABA,whereas for somatic embryo germination wasIBA, kinetin and GA 3 . The results show thatthe best transfer interval for callus proli-feration was four weeks. In this treatment therelative growth rate of callus was0.38 g/g/week. Somatic embryo initiation fromthe callus was done in DF mediumsupplemented with 1 mg/L 2,4-D and 0.1 mg/Lkinetin. The percentage of somatic embryowas 80% based on biomass fresh weight afterthe fourth subculture. The addition of 0.5 mg/Lkinetin and 0.05 mg/L ABA improved somaticembryo maturation of oil palm; the averagenumber of somatic embryos at advanced stages(torpedo and cotyledonary) was 16.3 embryosper flask. The addition of 2 mg/L IBA and0.5 mg/L kinetin in DF medium with half-strength macro-salt enhanced significantly thegermi-nation of somatic embryos. GA 3 at0.1 mg/L increased the total number ofgerminants.RingkasanPada sistem perendaman sesaat (SPS),bahan tanam hanya terpapar sebentar dalammedium sehingga paparan dengan udara lebihlama dan kekurangan oksigen yang seringterjadi pada kultur cair dapat diatasi. Penelitianini bertujuan menetapkan prosedur untukperbanyakan kalus embriogenik sampai denganperkecambahan embrio somatik kelapa sawit(Elaeis guineensis Jacq.) dalam kultur SPS.Kalus embriogenik kelapa sawit klon MK 638yang diperoleh dari Balai Penelitian Marihatdiperbanyak pada medium padat di ruang gelapyang kemudian digunakan sebagai bahan untukkultur cair SPS. Lama perendaman semuakultur di SPS diatur tiga menit denganfrekuensi setiap enam jam. Perbanyakan kalusdalam medium cair DF dengan 2,4-D 5 mg/Ldan kinetin 0,1 mg/L dilaksanakan denganinterval subkultur 4, 6 dan 8 minggu.Perlakuan pematangan embrio somatik adalahkinetin dan ABA sedangkan perlakuan untukperkecambahan embrio somatik adalah IBA,kinetin dan GA 3 . Hasil penelitian menunjuk-kan bahwa untuk proliferasi kalus embriogenikkelapa sawit, interval subkultur terbaik adalahempat minggu. Pada perlakuan ini laju tumbuhrelatif kalus mencapai 0,38 g/g/minggu.Inisiasi embrio somatik dari kalus dilakukanpada medium DF ditambah 2,4-D 1 mg/L dankinetin 0,1 mg/L. Persentase embrio somatikmencapai 80% dari total bobot basah biomassasetelah subkultur keempat. Penambahan kinetin0,5 mg/L dan ABA 0,05 mg/L meningkatkanpematangan embrio somatik kelapa sawit; rata-rata jumlah embrio somatik fase lanjut (torpedodan kotiledon) adalah 16,3 embrio per bejana.Penambahan IBA 2 mg/L dan kinetin 0,5 mg/Lpada medium DF dengan setengah garammakro meningkatkan perkecambahan embriosomatik secara nyata. GA 3 0,1 mg/L mening-katkan jumlah kecambah yang terbentuk.

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Morphological Characterization on Abnormalities of Oilpalm (Elaeis guineensis Jacq) Embryo Somatic Generated from Leaf Explant. Nesti F. Sianipar, Gustav A. Wattimena, Hajrial Aswidinnoor, Maggy Thenawidjaya S., Nurita Toruan-Mathius, and Gale Ginting. Somatic embryogenesis is the development of somatic cells to form a structure alike zygotic embryo direct or indirectly. Somatic embryos from young leaf explants could be induced from primary callus formed surrounding the palm-leaf rib. Embryogenic callus will develop to be somatic embryos which grew nonuniformly. Embryo somatic growth pattern of globular, asymmetric heart shape, and cotyledon ary stage produced different morphological variation. Morphological variability of in vitro somatic embryos could be due to high application of growth regulator 2,4-D at the beginning of initiation, subculture frequency, loaded cells, and polysomic cells from certain tissues. From the three clones used, which were clone 638, 636, and 558, there were different variation at each step of development stages, grouping morphologically into normal and abnormal based on the development of somatic embryos. The percentage of abnormality from the three clone used was clone 27% (638), 30% (636), and 46% (558). The normal somatic embryos at globular stage were round and bipolar shaped; while the abnormal embryos were oval and no bipolar. At heart-shape stage, the normal somatic embryos had symmetrical polarized surface; while the abnormal embryos had asymmetrical polarized surface. At the cotyledon stage, the normal embryos had monocot-tyledon; the abnormal ones were more than one cotyledon.

Embryo somatic (ES) abnormalities of  oil palm were probably caused by numbers and location of  DNA genom cytosin methylation. Quantity of methylation could be determined  by Reverse phase HPLC (RP-HPLC) techniques, while location of DNA cytosin methylation was detected by Random Amplified Fingerprint DNA (RAF-DNA) technique. The objective of  this research was to determine numbers  and pattern of DNA cytosin methylation  of normal or abnormal ES and normal ortet as a standard.  DNA genomic of samples were cut by  HpaII and MspI enzymes at  CCGG site, and amplified by RAF. HpaII  cut at  mCCGG  sequences, but if  second C  were methylated  the sequences can not be cut by HpaII.While Msp1 will cut if internal of cytosine was methylated  (CmCGG). The results showed that AB16, AE11, AO12 and  AP12 primers could detect the changes of methylation site on normal ortet and abnormal ES cotyledone.  RP-HPLC analyses showed that DNA cytosin methylation content between ES globular and ES cotyledone, both normal and abnormal  and also normal plantlet and ortet were unsignificantly different. DNA methyl content was around   0.25 – 2.72 %. Internal and fully methylation was found on 124 – 457 bp. Abnormal ES  of MK638 clone showed the hipomethylation pattern. It was concluded that methylation cytosine content was very low and it seems that DNA methylation undirectly  affects on process of morphology abnormalities.  Abnormalities of ES globular and cotyledone might be caused by the change of  DNA genom sequences. Abstrak Abnormalitas pada embrio somatik (ES) tanaman kelapa sawit diduga disebabkan oleh kandungan serta lokasi terjadinya metilasi sitosin DNA genom. Kandungan metilasi dapat ditetapkan dengan teknik Reverse Phase HPLC (RP-HPLC), sedang lokasi terjadinya  metilasi sitosin DNA genom ES dapat dideteksi dengan teknik Random Amplified Fingerprint DNA (RAF-DNA). Tujuan penelitian ini adalah untuk menetapkan pola metilasi sitosin DNA genom ES kotiledon normal dan abnormal, sebagai pembanding adalah ortet yang normal.  DNA genom contoh dipotong dengan enzim HpaII dan MspI yang mengenali situs CCGG, selanjutnya diamplifikasi dengan RAF. Enzim HpaII memotong sekuen mCCGG tetapi jika C kedua mengalami metilasi sekuen tersebut tidak terpotong. Msp1 akan memotong apabila sitosin internal termetilasi (CmCGG). Hasil yang diperoleh menunjukkan bahwa terjadi perubahan  situs metilasi antara ortet  normal dan ES kotiledon abnormal. Perubahan situs metilasi sitosin dapat dibedakan dengan primer RAF, yaitu AB16, AE11, AO12 dan AP12.  Hasil analisis RP-HPLC menunjukkan bahwa perbedaan kandungan metilasi sitosin DNA ES globular maupun kotiledon masing-masing antara yang normal dan abnormal, serta antara planlet dan ortet normal sangat kecil. Kandungan metil sitosin berkisar antara 0,25 – 2,72 %. Tampak bahwa pada 124 - 457 pb terjadi metilasi internal, eksternal maupun metilasi penuh. Pada ES abnormal klon MK638 menunjukkan terjadi hipometilasi sitosin.  Perbedaan kandungan  metilasi sitosin yang sangat kecil diduga tidak berpengaruh langsung terhadap proses abnormalitas. Abnormalitas yang terjadi pada ES globular dan kotiledon kemungkinan akibat terjadinya perubahan sekuens DNA genom.