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Characterization Of Zygotic And Nucellar Embryo Of Six Indonesian Mango Cultivars Using Molecular Markers Fatimah, Fatimah; Husni, Ali; Kosmiatin, Mia; Karsinah, Karsinah; Baroya, Mushlihatun
ANNALES BOGORIENSES Vol 20, No 2 (2016): Annales Bogorienses
Publisher : Research Center for Biotechnology - Indonesian Institute of Sciences (LIPI)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (458.762 KB) | DOI: 10.14203/ann.bogor.2016.v20.n2.69-75

Abstract

Breeding mangoes is a long term activity and complicated. One of difficulties is polyembryonic seeds. These factors make genetic improvement through conventional parental selection and breeding slow and unpredictable. Adoption of molecular markers and genomics-based breeding strategies will likely improve predictability and breeding efficiency. The objective of this study was to evaluate the occurrence of zygotic or nucellar in polyembryony mango cultivars Garifta Merah, Lalijiwo, Manalagi, Madu, Saigon Kuning, and Saigon Merah using RAPD and SSR markers. The type of embryo (zygotic or nucellar) was evaluated by comparing the exhibiting amplification patterns, if different from the mother plant considered as zygotic and identified as nucellar if they exhibited the same banding pattern as the mother plant. Out of the 16 SSR and 16 RAPD primers evaluated, 9 primers each amplified the largest number of allele and sharply defined band. Dendogram of the SSR primers divide into three groups while RAPD primers distinguish clearly into two major groups, zygotic group and nucellar group. Based on zygotic and nucellar size, number and position, indicating no relationship between the type of embryo and embryo size, number and the position. 
Molecular diversity of citrus genotypes using callose synthase 7 gene markers linked to Huanglongbing resistance Nugroho, Kristianto; Purwito, Agus; Sukma, Dewi; Kosmiatin, Mia; Santoso, Tri Joko; Husni, Ali; Martasari, Chaireni; Lestari, Puji
Jurnal Agronomi Indonesia (Indonesian Journal of Agronomy) Vol. 53 No. 2 (2025): Jurnal Agronomi Indonesia (Indonesian Journal of Agronomy)
Publisher : Indonesia Society of Agronomy (PERAGI) and Department of Agronomy and Horticulture, Faculty of Agriculture, IPB University, Bogor, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24831/jai.v53i2.64952

Abstract

Huanglongbing (HLB), caused by Candidatus Liberibacter asiaticus, is a notable disease affecting citrus plantations globally. Several studies showed that the callose synthase 7 gene is crucial for the citrus defense system against this pathogen. The study aimed to analyze the nucleotide variations and genetic diversity among several citrus genotypes using specific gene primers designed from the callose synthase 7 gene sequence. Genomic DNA from eleven citrus genotypes was amplified using the specific primers, and Sanger sequencing was employed to identify the nucleotide sequence of the PCR products. The results revealed a total of 66 single-nucleotide polymorphisms (SNPs), 10 insertions, and 11 deletions were detected in callose synthase 7 gene fragment sequences. Of these, one out of five noteworthy SNPs identified at a position of 200 bp downstream of the START codon showed distinguishing features between susceptible and resistant/tolerant genotypes. Phylogenetic analysis clearly discriminated the eleven citrus genotypes into two clusters at a dissimilarity coefficient of 0.05, with all genotypes grouped in the first cluster, except for the Chinese box orange and orange jasmine. The identification of notable SNPs in this study can aid in developing new markers for the rapid selection of genotypes with enhanced HLB resistance in citrus breeding programs.   Keywords: Candidatus Liberibacter asiaticus; nucleotide variations; Sanger sequencing; phylogenetic analysis; SNP
Nucleotide variations of WRKY70 gene sequence related to Huanglongbing resistance in citrus Nugroho, Kristianto; Purwito, Agus; Sukma, Dewi; Kosmiatin, Mia; Santoso, Tri Joko; Reflinur, Reflinur; Mastur, Mastur
Jurnal Biologi Tropis Vol. 25 No. 4 (2025): Oktober-Desember
Publisher : Biology Education Study Program, Faculty of Teacher Training and Education, University of Mataram, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.29303/jbt.v25i4.10092

Abstract

Huanglongbing, inflicted by Candidatus Liberibacter asiaticus in Asia region, is a destructive disease affecting citrus productions worlwide. Several studies have identified resistance genes that play essential roles in the citrus defense system against this pathogen. The goals of this study were to design the specific gene primers from the WRKY70 gene sequence and analyze the nucleotide variations and genetic diversity among several citrus genotypes. Genomic DNA from nine citrus genotypes were amplified using WRKY70-specific gene primers and the products of PCR were sent to Sanger sequencing, while the sequences of the other 12 genotypes were collected from Citrus Genome Database. The results revealed a total of 282 nucleotide variations which consisted of 157 SNPs, 28 insertions, and 97 deletions, were identified in the WRKY70 gene fragment sequence. There were three notable SNPs detected, with only one SNP [C/T] in first intron area at the position of 524 bp downstream from START codon that showed its ability to distinguish between susceptible and tolerant/resistant citrus genotypes. The phylogenetic analysis also revealed the clearly separation among citrus genotypes in two main clusters. The discovery of this SNP is useful for designing a functional marker as a screening tool in citrus breeding program in the future.
Perkecambahan dan Pertumbuhan in vitro Jeruk Nipis (Citrus aurantifolia) Komang Trisna Wirakusuma; Purwito, Agus; Husni, Ali; Kosmiatin, Mia
Jurnal Hortikultura Indonesia (JHI) Vol. 14 No. 1 (2023): Jurnal Hortikultura Indonesia
Publisher : Indonesian Society for Horticulture / Department of Agronomy and Horticulture

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.29244/jhi.14.1.1-8

Abstract

In vitro selection to increase the citrus resistance to Huanglongbin-HLB diseases, requires theavailability of lime in vitro shoots as a negative control, because it is one type of citrus that is verysusceptible to HLB. In vitro shoots can be obtained from seed germination in vitro with a more controlledenvironment. Shoots derived from in vitro germination can be used as controls when in vitro artificialinoculation by pathogen suspension is performed to select in vitro breeding lines. This study aims to obtainthe best concentration of gibberellins and scarification treatment for in vitro germination of lime.Germination was carried out on citrus seeds that had been sterilized on the surface of the fruit. Scarificationis done by injuring the testa of the seed. Lime seeds with and without scarification treatment were germinated on MS+VMW base media with the addition of GA3 0 mg L-1, 5 mg L-1, 10 mg L-1, and 15 mgL-1. The results showed that the optimal concentration of GA3 to induce lime seed germination was 5 mgL-1. Scarification by wounding the seed testas significantly accelerated lime germination time. Keywords: Citrus aurantifolia, in vitro selection, negative control, of GA3, scarification
Karakterisasi dan Kelayakan Ekonomi Citrus aurantifolia kultivar ‘Nimas Agrihorti’ sebagai Komoditas Biofarmasi Potensial di Indonesia Budiyati, Emi; Waluyo, Budi; Maghfoer, Moch. Dawam; Kosmiatin, Mia; Triwiratno, Anang; Hanif, Zainuri; Andrini , Anis; Hanim, Faridha; Tina , Siti Agus
Buitenzorg: Journal of Tropical Science Vol 2 No 1 (2025): Vol. 2 No. 1 (2025): Buitenzorg: Journal of Tropical Science
Publisher : Innovation Centre for Tropical Sciences

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.70158/buitenzorg.v2i1.17

Abstract

‘Nimas Agrihorti’ is a citrus cultivar developed to fulfil the increasing domestic demand for biopharmaceutical-grade citrus in Indonesia. Its development aligns with the growing public interest in health-promoting agricultural products. This study aimed to evaluate the morphological characteristics, chemical composition, and economic viability of ‘Nimas Agrihorti’ as a biopharmaceutical citrus. Field observations were conducted from June 2019 to October 2020 at the Banjarsari Experimental Field, Bayeman Village, Probolinggo, East Java. Morphological assessments followed the Descriptor List for Citrus by IPGRI, and chemical analyses were performed at the Post-Harvest Laboratory of Brawijaya University, Malang. Economic feasibility was assessed by calculating production costs and revenues over a 2-year cultivation cycle. ‘Nimas Agrihorti’ exhibited favorable morphological traits, including large fruit size (72–82 g), yellow skin, and sweet taste. Chemical analysis revealed vitamin C content of 34.8 mg/100 g and low acidity at 0.45%. The total production cost over 2 years was IDR 42,640,000.00 (equivalent to USD 2,584), with total revenue amounting to IDR 77,360,000.00 (equivalent to USD 4,688), resulting in a Revenue-Cost (R/C) ratio of 2.81. The combination of desirable morphological and chemical characteristics, along with strong economic returns, indicates that ‘Nimas Agrihorti’ holds significant potential for development as a biopharmaceutical citrus cultivar in Indonesia.   Keywords: citrus, cultivar, biopharmaceutical products, economic feasibility
Induksi Embriogenesis Somatik dari Jaringan Endosperma Jeruk Siam (Citrus nobilis Lour.) cv Simadu Kosmiatin, Mia; Purwito, Agus; Wattimena, Gustaff Adolf; Mariska, Ika
Jurnal Agronomi Indonesia (Indonesian Journal of Agronomy) Vol. 42 No. 1 (2014): Jurnal Agronomi Indonesia
Publisher : Indonesia Society of Agronomy (PERAGI) and Department of Agronomy and Horticulture, Faculty of Agriculture, IPB University, Bogor, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (610.396 KB) | DOI: 10.24831/jai.v42i1.8149

Abstract

ABSTRACTTriploid plants can be obtained from endosperm tissues through somatic embryogenesis regeneration. This research aimed to obtain somatic embryogenesis regeneration technique of tangerine endosperm. There were 3 experiments conducted in this research: 1) Embryogenic callus induction of tangerine endosperm. Endosperms isolated from fruits that were harvested from mother plants 11-13 weeks after anthesis and cultured on Murashige and Skoog (MS) medium by modified vitamin Morel and Wetmore (MW) which treated by 0.1 mg L-1 biotin, 500 mg L-1 malt extract (ME), 500 mg L-1 Casein hydrolisate (CH), 500 mg L-1 ME + 0.1 mg L-1 biotin, and 500 mg L-1CH + 0.1 mg L-1 biotin, 2) Maturation and germination of somatic embryos conducted by embryogenic callus cultured on MS medium by vitamin MW modified with addition of ABA, glutamine, and biotin, and 3) Plantlet elongation conducted on MS medium modified by MW vitamin with addition of GA3 and Kinetin. The best induction medium for embryogenic callus was modified MS enriched with 3 mg L-1 BA and 500 L-1 CH or ME, in succession 84.0 and 80.0%. The best medium for somatic embryos maturation with normal morphological plantlets (54.8%) was modified MS medium without plant growth regulator with higher rate of solidified agent (from 2.5 to 3 g L-1 Phytagel). Plantlets elongation was highly (0.9 cm) occurred on modified MS with enriched of 2.5 mg L-1 GA3. Keywords: Citrus nobilis (Lour.), endosperm culture, in vitro, Simadu tangerine
Transformation of Ponkan Mandarin (Citrus reticulata Blanco) by CRISPR/Cas-9-gRNA-CsCS to increase plant resistance to huanglongbing disease Dian Puji Rahayu; Purwito, Agus; Kosmiatin, Mia; Husni, Ali
Jurnal Agronomi Indonesia (Indonesian Journal of Agronomy) Vol. 52 No. 2 (2024): Jurnal Agronomi Indonesia (Indonesian Journal of Agronomy)
Publisher : Indonesia Society of Agronomy (PERAGI) and Department of Agronomy and Horticulture, Faculty of Agriculture, IPB University, Bogor, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24831/jai.v52i2.53465

Abstract

The gene that regulates callose could be modified by CRISPR/Cas9 technology. This research aimed to insert the CRISPR/Cas9-CsCS gene into Ponkan orange genome using Agrobacterium tumefaciens. The explants were soaked in a bacterial suspension for 20 minutes and incubated for 2-3 days. In vitro acceleration growth was conducted with a two-factor completely randomized design. The first factor is the type of explant with three levels (nucellar embryo, zygotic embryo, cotyledon node), and the second factor is the type of media with 6 levels (VMW, MT, MSK0, MSK1, MSK2, MSK3). The results showed of all explant types, the highest plant height average and number of leaves were obtained in a media combination of MS + Kinetin 2 mg L-1, MT, and MS + Kinetin 3 mg L-1. The highest transformation efficiency was in the nucellar embryo explant, while the highest regeneration efficiency was in the zygotic embryo explant. The highest shoot tip grafting percentage was achieved in the cotyledon node explant at 100%. In the grafting phase, the putative transformants before and after artificial bacterial inoculation showed that Ponkan 606 and Ponkan 597 had the highest plant heights, respectively. The intensity of Huanglongbing attacks after bacterial inoculation showed that three genotypes did not show HLB symptoms in the 24th week of observation, namely genotypes 598, 606, and 607. This study concluded that gene transformation in citrus plants produced three genotypes that did not show HLB symptoms. Keywords: transgenic; Agrobacterium tumefaciens; Citrus Vein Phloem Degeneration (CVPD)
OPTIMIZATION OF KINETIN CONCENTRATIONS AND MEDIUM COMPOSITIONS FOR CITRUS SHOOT MULTIPLICATION FROM COTILEDONARY NODE Nugroho, Kristianto; Kosmiatin, Mia; Santoso, Tri Joko; Sukma, Dewi; Purwito, Agus; Husni, Ali; Martasari, Chaireni
BIOTROPIA Vol. 31 No. 1 (2024): BIOTROPIA Vol. 31 No. 1 April 2024
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.11598/btb.2024.31.1.2136

Abstract

Plant regeneration post-genetic transformation play an important an role in genome editing activities that should be optimized via several factors such as the composition of the medium and the concentration of plant growth regulators. This study aimed to optimize the kinetin concentrations and medium compositions for shoot multiplication originating from cotyledon node explants of several local citrus cultivars. The cotyledonary nodes from three citrus cultivars (Batu 55, Siam Madu, and Proksi-1 Agrihorti) were incubated in MS medium with Morel and Wetmore vitamins (VMW) supplemented with several kinetin concentrations (0; 0.2; 0.4; 0.6; 0.8; and 1 mg/L). The best kinetin concentrations for number of shoots variable were then combined with MT medium. The results showed that kinetin concentration at 0.8 mg/L gave the best number of shoot in Batu 55 cultivar as well as 1 mg/L concentration in Siam Madu and Proksi-1 Agrihorti cultivars. The combination 1 mg/L kinetin with Murashige and Tucker (MT) medium showed the best number of shoots, percentage of shoot formation, number of leaves, number of nodes, and shoot length in this study. This medium composition could be further used for shoot multiplication in genetic transformation in those three citrus cultivars, including genome editing activities in development of new improved citrus varieties.
Characterization of Zygotic and Nucellar Embryo of Six Indonesian Mango Cultivars Using Molecular Markers Fatimah, Fatimah; Husni, Ali; Kosmiatin, Mia; Karsinah, Karsinah; Baroya, Mushlihatun
Annales Bogorienses Vol. 20 No. 2 (2016): Annales Bogorienses
Publisher : BRIN

Show Abstract | Download Original | Original Source | Check in Google Scholar

Abstract

One of difficulties in mangoes hybridization is polyembryonic seeds. This phenomenon reduces the chance of recovering true hybrid seedlings. The identification of the zygotic embryo is difficult and the possible approach is by using molecular marker. The objective of this study was to evaluate the utilization of two different marker systems (dominant markers and SSR) to characterize the occurrence of zygotic or nucellar embryo in polyembryony mango cultivars Garifta Merah, Lalijiwo, Manalagi, Madu, Saigon Kuning, and Saigon Merah. The type of embryo was evaluated by comparing the exhibiting amplification patterns, if different from the mother plant considered as zygotic and identified as nucellar if they exhibited the same banding pattern as the mother plant. From both of this marker systems out of the 16 SSR and 16 evaluated dominant primers, nine primers of each systems amplified the largest number of allele and sharply defined band. Dendogram analysis showed that the evaluated dominant markers could distinguish the zygotic and nucellar embryo clearly compare to evaluated SSR primers. The number of zygotic embryos derived from SSR was 64% and dominant markers were 47%. Based on zygotic and nucellar size, number and position, indicating no relationship between the type of embryo and embryo size, number and the position.