Abstract. Maryanto SD, Tanjung ZA, Roberdi, Sudania WM, Pujianto, Hairinsyah, Utomo C, Liwang T. 2021. Involvement of purple acid phosphatase gene into nitrogen uptake of oil palm (Elaeis guineensis). Biodiversitas 22: 1385-1390. Nitrogen is the most important nutrient element in terms of plant growth. Plant purple acid phosphatases (PAPs) are known to participate in the phosphate (Pi) acquisition and utilization. Moreover, PAP gene plays an important role in nitrogen fixation. A single nucleotide polymorphism (SNP) was previously detected in the exon 7 of EgPAP3, based on SNP mining analysis of oil palm (Elaeis guineensis Jacq.). genome database. This study was aimed to obtain a Cleaved Amplified Polymorphic Sequences (CAPS) marker based on SNP within EgPAP associated with efficient nitrogen uptake oil palm progenies. Primer pairs were designed and used for PCR amplification of 3 oil palm progenies that showed low N-content, 3 progenies with moderate N-content, and 3 progenies with high N-content. The amplicon was purified prior to single-pass DNA sequencing analysis. Based on Pearson’s chi-square and odds ratio statistical analysis, the SNP has strong positive correlation with the phenotype. The SNP is located at chromosome 13 with a distance of 17.7771 cM from start codon. The sequencing analysis revealed that three progenies with high N-content samples had GG allele motif, while moderate N-content progenies had GA allele and low N-content progenies had AA allele motifs respectively. In addition, a restriction site of NIaIV was found to be adjacent to the SNP location, thus the PCR products of all samples were digested with NIaIV restriction enzyme. NIaIV was able to distinguish between high, medium and low efficient DNA samples. Whole high N-content progenies with GG allele motifs were undigested indicating a single band size of 670 bp identical to the untreated PCR product (control). Moderate N-content progenies produced a 670 bp, 550 bp, and 120 bp bands because of digested by NIaIV. Low N-content progenies also resulted in double bands of 550 bp and 120 bp due to digested by NIaIV. Furthermore, NIaIV restriction enzyme was applied to digest other 54 oil palms DNA samples with unknown genotypes. Whole GG samples were consistently shown to have single band, GA and AA samples were also consistent in producing two bands with different lengths. Based on this result, CAPS marker based on SNP in EgPAP3 was successfully developed to screen between high and low efficient N-uptake of oil palm progenies.

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Abstract. Saputra TI, Roberdi, Nugroho YA, Artutiningsih W, Purba OS, Maryanto SD, Yono D, Utomo C, Liwang T. 2021. The development of unlabeled probes-high resolution melting (UP-HRM) marker on SAD, IAA27 and ACC genes of oil palm. Biodiversitas 22: 3356-3362. The unlabeled probes-high resolution melting (UP-HRM) marker is a useful tool for detecting of single nucleotide polymorphisms (SNPs). The objectives of this study were to develop UP-HRM markers to differentiate specific SNPs patterns on oil palm. The marker was developed and tested with Elaeis guineensis (Eg), Elaeis oleifera (Eo), Eo x Eg (hybrid), and was validated with 53 individuals of BC1F1 populations ((Eo x Eg) x Eg). Four UP-HRM markers were developed based on 2 SNPs in the stearoyl-acyl-carrier-protein 9-desaturase (EgSAD), 1 SNP in the auxin-responsive protein IAA27-like (EgIAA27), and 1 SNP in the 1-amino cyclopropane-1-carboxylate oxidase (EgACC) genes. The SNP discovery result showed that Eg was represented a reference homozygote genotype, while Eo was represented as an alternative homozygote genotype and the Eo x Eg hybrid was represented as a heterozygote genotype in all genes. The typical UP-HRM melt curve graph was successfully generated. This result was consistent with each genotype model for all four markers. The UP-HRM markers can distinguish each genotype according to the single-pass sequencing results. Furthermore, dendrogram analysis on validation divided 53 BC1F1 samples into three cluster groups.

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Optimisation of Single Nucleotide Amplified Polymorphism (SNAP) Primers in Brassinosteroid (BRI) Gene of Oil Palm Oil palm varieties that have high yields still have a high rate of stem height increase, and it has an impact on difficulties at harvest. Brassinosteroid (BRI) is one of the growth hormones that regulate stem height increase. In this study, the sample used is three types of oil palm, namely E. guineensis which represents the height growth of normal oil palm stems, E. oleifera which represents slow stem height growth, and the result of crossing or hybrid Eo x Eg which is expected to inherit the traits between the two the crossed parent. Sequencing was carried out on samples using a specific primer for the BRI gene that triggers elongation and division of stem cells. The Single Nucleotide Amplified Polymorphisms (SNAP) primers used in the analysis process need to find the optimum temperature to obtain the optimum PCR conditions. Four primers of Single Nucleotide Amplified Polymorphism (SNAP) that get the optimum temperature with a mixture of normal reagents are BRI 69 Alt-Rev and BRI 562 Ref-Rev at 55oC, BRI 1206 Alt-Rev at 57oC and BRI 1206 Ref-Rev at 58oC. Two primers require additional MgCl2 in the reactant mixture, namely BRI 69 Ref-Rev at 55oC and BRI 2115 Alt-Rev at 56oC.Keywords: Oil palm, Brassinosteroid, Primer, DNA, GenesABSTRAK Varietas-varietas kelapa sawit yang memiliki daya hasil tinggi umumnya masih memiliki laju pertambahan tinggi yang relatif cepat dan berakibat pada kesulitan saat panen. Brassinosteroid (BRI) merupakan salah satu hormon pertumbuhan yang mengatur pertambahan tinggi batang. Pada studi ini, Sampel yang digunakan adalah tiga jenis kelapa sawit yaitu sampel E. guineensis yang mewakili pertumbuhan tinggi batang kelapa sawit normal, E. oleifera  yang mewakili pertumbuhan tinggi batang lambat, dan hasil persilangan atau hibrida Eo x Eg yang diharapkan mewarisi sifat antara kedua induk yang disilangkan. Sekuensing dilakukan pada sampel menggunakan primer spesifik gen BRI yang memicu pemanjangan dan pembelahan sel batang. Primer Single Nucleotide Amplified Polymorphisms (SNAP) yang digunakan dalam proses analisis perlu dicari suhu optimumnya agar didapatkan kondisi PCR yang optimum sehingga dihasilkan produk PCR yang spesifik. Empat primer Single Nucleotide Amplified Polymorphism (SNAP) yang mendapatkan suhu optimum dengan campuran pereaksi normal yaitu BRI 69 Alt-Rev dan BRI 562 Ref-Rev pada suhu 55oC, BRI 1206 Alt-Rev pada suhu 57oC dan BRI 1206 Ref-Rev pada suhu 58oC. Dua primer membutuhkan tambahan MgCl2 pada campuran pereaksinya yaitu BRI 69 Ref-Rev pada suhu 55oC dan BRI 2115 Alt-Rev pada suhu 56oC.Kata Kunci: Kelapa sawit, Brassinosteroid, Primer, DNA, Gen

Oil palm micropropagation through tissue culture is a technique to provide elite oil palms to meet the desired traits. This technique is commonly carried out through somatic embryogenesis. However, the oil palm’s somatic embryogenesis rate is quite low. Several approaches have been made to overcome this problem, including transcriptome profiling through RNA-seq to identify key genes involved in oil palm somatic embryogenesis. RNA sequencing was applied in high- and low-embryogenic ortets of Tenera varieties based on the somatic embryoid rate at the callus, globular, scutellar, and coleoptilar embryoid stages. Cellular analysis of embryoid inductions and proliferations showed that high-embryogenic ortets resulted in higher embryoid proliferation and germinations than low-embryogenic ortets. Transcriptome profiling showed that there are a total of 1,911 differentially expressed genes (DEGs) between high- and low-embryogenic ortets. ABA signaling-related genes such as LEA, DDX28, and vicilin-like protein are upregulated in high-embryogenic ortets. Furthermore, DEGs associated with other hormone signaling, such as HD-ZIP associated with brassinosteroids and NPF associated with auxin, are upregulated in high-embryogenic ortets. This result suggests a physiological difference between high- and low-embryogenic ortets that is connected to their capacity for somatic embryogenesis. These DEGs will be used as potential biomarkers for high-embryogenic ortets and will be validated in further studies.

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