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Molecular Characterization of a Rigid Rod-Shaped Virus Isolated from Frangipani (Plumeria sp.) Showing Mosaic Symptom in Taiwan Choliq, Fery Abdul; Chen, Tsang Hai; Sulistyowati, Liliek
Journal of Tropical Plant Protection Vol 1, No 1 (2012)
Publisher : University of Brawijaya

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

Frangipani is an important succulent plant around the worlds and also in Taiwan, for example, Plumeria rubra is widely grown as a popular ornamental tree in parks and landscaped establishments in Taiwan. Recently, a new disease in frangipani with mosaic and distortion symptoms was found in Taiwan. No viruses caused frangipani disease has been reported in Taiwan and the references about frangipani disease are still limited and only Frangipani mosaic virus (FrMV) was found. In this study, the molecular properties of a virus isolated from symptomatic frangipani in south Taiwan, such as Pingtung, Kauhsiung and Tainan were investigated. The virus with rod-shaped particles of 300 nm long and 18 nm in diameter was examined inside diseased leaves by electron microscopy. The purified virus particles showed the typical UV spectrum of tobamoviruses with A260/A280 value of 1.29 and maximum and minimum absorption at 260 nm and 249 nm, respectively. The molecular weight of19.5 kDa as the size of coat protein of tobamoviruses was estimated bysodium dedocyl sulfate-polyacrylamide gel (SDS-PAGE). Furthermore, the degenerate primers for tobamoviruses were used to amplify 568 bp and 400 bp of the DNA fragments in RT-PCR and nested PCR, respectively. Based on these results, it was confirmed that the rigidrod-shaped virus isolated from mosaic symptom of frangipani leaves is an isolate of FrMV, belonging to the genus Tobamovirus. This is the first report thatFrMV infecting Plumeria sp.in Taiwan.Â Keywords: Frangipani plant, mosaic disease, FrMV, Tobamovirus

TRANSFER GEN   -1,3-GLUCANASE DARI JAMUR Trichoderma asperillum PADA KALUS ABAKA UNTUK KETAHANAN TERHADAP PENYAKIT LAYU FUSARIUM RULLY DYAH PURWATI; LILIEK SULISTYOWATI
Jurnal Penelitian Tanaman Industri Vol 18, No 1 (2012): Maret 2012
Publisher : Pusat Penelitian dan Pengembangan Perkebunan

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.21082/jlittri.v18n1.2012.24-30

ABSTRAKKendala utama dalam budidaya tanaman abaka (Musa textilis Nee.)adalah penyakit layu Fusarium yang disebabkan oleh Fusarium oxysporumf.sp cubense (Foc). Upaya perbaikan sifat ketahanan tanaman abakamelalui persilangan sulit dilakukan karena keragaman genetiknya sempitakibat pola perbanyakan secara vegetatif yang terus-menerus.Transformasi gen ketahanan β-1,3-Glucanase merupakan salah satualternatif untuk memperbaiki sifat ketahanan tanaman dengan bantuanvektor Agrobacterium tumefaciens. Gen  -1,3-Glucanase diisolasi darijamur endofit Trichoderma asperillum yang diketahui antagonis terhadapFusarium oxisporum. Penelitian bertujuan untuk mengintroduksi gen β-1,3 Glucanase pada tanaman abaka, sebagai tahap awal untuk memperolehtanaman abaka tahan terhadap penyakit layu Fusarium. Penelitiandilaksanakan di Laboratorium Bioteknologi Fakultas Pertanian, Univer-sitas Brawijaya dan Laboratorium Kultur Jaringan Balai PenelitianTanaman Tembakau dan Serat, mulai Juni 2007 sampai dengan Mei 2009.Penelitian terdiri atas tiga tahap sebagai berikut: transfer gen  -1,3-Glucanase pada kalus abaka embriogenik, regenerasi tunas dan planletabaka transforman, dan konfirmasi planlet abaka transforman yangmengandung gen Gus dan  -1,3-Glucanase. Transfer gen dilakukanmelalui vektor A. tumefaciens strain LBA4404 yang mengandung plasmidpB2GW7 berisi gen-gen  -1,3-Glucanase, Gus (  -glucuronidase) sebagaigen pelapor dan Bar (Basta resistance) sebagai gen penyeleksi. Kalusabaka klon UB-13 embriogenik berukuran 3 x 3 x 3 mm 3 direndam dalamsuspensi A. tumefaciens, kemudian ditanam pada media kokultivasi selama2 hari. Setelah kokultivasi, kalus dipindahkan ke media MS cair+Timentin100 ppm selama 2 minggu. Selanjutnya kalus dipindahkan ke mediainduksi kalus (MK) yaitu MS + BAP 5 mg/l + Thidiazuron 0,4 mg/l +vitamin C 100 mg/l + Basta 50 ppm + Timentin 100 ppm. Regenerasitunas dilakukan dengan memindahkan kalus transforman ke media induksitunas (MT): MS+BAP 0,5 mg/l + vitamin C 100 mg/l dengan penambahandan tanpa Timentin 100 ppm. Tunas transforman dengan tinggi 2-3 cmdipindahkan ke dalam media induksi akar (MA) : MS + arang aktif 2 g/ldengan penambahan dan tanpa Timentin 50 ppm. Keberadaan gen Gusdideteksi dengan reaksi histokimia, dan konfirmasi keberadaan gen  -1,3-Glucanase dilakukan dengan Polymerase Chain Reaction (PCR). Daripenelitian berhasil diperoleh 2% kalus transforman yang lolos seleksiBasta. Hasil konfirmasi keberadaan gen Gus pada planlet transformanmenunjukkan 9 dari 20 (45%) planlet yang diuji, positif mengandung genGus. Konfirmasi keberadaan gen  -1,3-Glucanase dengan PCRmenunjukkan hanya 2 dari 20 planlet transforman, positif mengandung  -1,3-Glucanase. Pengujian ketahanan dari plantlet transgenik tersebut perludilakukan terhadap Fusarium oxisporum f.sp cubense (Foc).Kata kunci: Musa textilis Nee., transformasi gen,  -1,3-Glucanase,Agrobacterium tumefaciens, penyakit, jamur patogen,FusariumABSTRACTThe main constraint of abaca (Musa textilis Nee.) cultivation isinfection of wilt disease caused by Fusarium oxysporum f.sp cubense(Foc). The effort to improve abaca resistance through hybridization is stilldifficult due to narrow genetic variability resulted from continuousvegetative multiplication. Transformation of  -1,3-Glucanase resistancegene is an alternative way to improve character of genetic resistance withhelp of Agrobacterium oxisporum. The research aimed at introducing  -1,3-Glucanase gene to abaca plants prior to obtaining the plants resistanceagainst Fusarium wilt diseases. The research was conducted inBiotechnology Laboratory, Faculty of Agriculture Brawijaya Universityand Tissue Culture Laboratory of Indonesian Tobacco and Fibre CropsResearch Institute, from June 2007 to May 2009. This experimentconsisted of three steps, namely:  -1,3-Glucanase gene transfer onto abacaembriogenic calli, regeneration of transgene abaca shoots and plantlets,and confirmation of transgene abaca plantlets containing Gus and  -1,3-Glucanase genes. Gene transfer was performed using A. tumefaciensvector strain LBA4404 with pB2GW7 containing genes of  -1,3-Glucanase and Gus (  -glucuronidase) as reporter, and Bar (Bastaresistance) as selector marker. Embriogenic calli of abaca clone UB-13were soaked in A. tumefaciens suspension and then cultured in co-cultivation medium for two days. After co-cultivation, calli weretransferred to liquid of MS medium + 100 ppm Timentine for two weeks.Furthermore, the calli were sub-cultured into callus induction medium :MS + 5 mg BAP/l + 0.4 mg Thidiazuron/l + 100 mg vitamin C/l + 50 ppmBasta + 100 ppm Timentine. Shoots regeneration was conducted bytransferring transgene calli to shoot induction medium : MS + 0.5 mg/lBAP + 100 mg vitamin C/l with and without addition of 100 ppmTimentine. Transgene shoots with 2-3 cm height were sub-cultured to rootinduction medium : MS + 2 g active charcoal/l with and without additionof 50 ppm Timentine. Detection of Gus gene was conducted usinghistochemical reaction, while confirmation of  -1,3-Glucanase gene wasperformed by PCR. This project resulted in 2% transgene calli passingBasta selection. Nine out of 20 plantlets (45%) confirmed the existance ofGus gene. PCR results showed that only 2 out of 20 transformed plantlets positively contained  -1,3-Glucanase gene. The plantlets resistanceagainst Fusarium oxisporum f.sp cubense (Foc) needs to be evaluated.Key words: Musa textilis Nee, gene transformation,  -1,3-Glucanase,Agrobacterium tumefaciens, plant disease, fungal disease,Fusarium

Biology and Predatory Behavior of Metioche vittaticollis (Stal) (Orthoptera: Gryllidae) Sri Karindah; Bagyo Yanuwiadi; Liliek Sulistyowati
Journal of Tropical Plant Protection Vol 1, No 1 (2012)
Publisher : University of Brawijaya

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (237.956 KB)

Metioche vittaticollis (Stal) is one of the generalist predator in rice field habitat. The biology and predatory behavior were studied in the laboratory. The life cycle of M. vittaticollis (Stal) averaged 40–61 days at 26°-28oC. The eggs were inserted singly within the leaf sheath of rice or weeds and hatched in 14.28 days. The nymphal period was varied between 27 and 45 days and passed four nymphal stadia. Female fecundity was averaged 50 eggs during her lifetime. The longevity of the female or male adult was ranged from 20 to 38 days. The average longevity of females and males were 29.24 and 25.00 days, respectively. The longevity of unmated female or male were longer than the mated female or male. The egg and first instar nymph sustained the high mortality of 30% and 25%, respectively, whilst there was less mortality in the third and fourth instar nymph. The adult females of M. vittaticollis survived for 32 days and the rate of survival was high in the young adults but decreased as the cricket aged. The females were more preferred to Brown Plant Hopper (BHP) nymph than the males. The early nymph stage of prey was the most stage to be fed by M. vittaticollis. However, the predation declined when they were given prey of late instar of BPH nymphs. Fewer adult stage of BPH was consumed by both male and female crickets.Key words: biology; generalist predator; prey consumption; Metioche vittaticollis

Hubungan Antara Aktivitas Poligalakturonase dengan Virulensi RAS 4 Fusarium oxysporum f.sp. cubense Arif Wibowo; Siti Subandiyah; Christanti Sumardiyono; Liliek Sulistyowati; Peter Taylor
Jurnal Perlindungan Tanaman Indonesia Vol 14, No 1 (2008)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.11871

One of the major constraints of banana plantation in Indonesia is the occurrence of fusarium wilt disease caused by Fusarium oxysporum f.sp. cubense. The pathogen produced series of cell wall degradation extracellular enzymes which have important roles in pathogenicity. Many studies have been conducted to know the role of degrading enzyme banana pectin is the major component of cell wall. Many pectinolytic enzymes such as polygalacturonase and others have been isolated from many fungal plant pathogens. The study was aimed to know the role of polygalacturonase towards the virulence of race 4 of F. oxysporum f.sp. cubense. The result showed that from 10 isolates of race 4 of F. oxysporum f.sp. cubense, the most virulent isolate was Lmp1 followed by Srg1, Bgl6, Mln1, Bgl3, A13, Bnt2, Gnk3, Kjg1 dan Wsb5. This was indicated by high and low percentage of wilting leaves of banana cultivar Cavendish when they were inoculated with these isolates. Incubation period varied from 3 to 6 weeks after inoculation SDS-PAGE showed that polygalacturonase, mostly PG1 and PG2, was secreted by these isolates, whereas PG3 was only found in growing cultures of Gnk3 and Wsb5 isolates. Detection of polygalacturonase activity with diffusion agar and reducing sugar methods showed that the activity of polygalacturonase secreted by F. oxysporum f.sp. cubense in the growing culture had no correlation with the virulence of the fungal pathogen.

Developments of Rice Cell Suspension Culture and A Novel Strategy for Screening New Resistant Lines to Rice Blight Disease Caused by Xanthomonas oryzae pv. oryzae Restu Rizkyta Kusuma; Liliek Sulistyowati; Chiu-Chsiung Cheng; Yi-Hsien Lin
AGRIVITA, Journal of Agricultural Science Vol 40, No 3 (2018): OCTOBER
Publisher : Faculty of Agriculture University of Brawijaya in collaboration with PERAGI

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.17503/agrivita.v40i3.1779

The research aimed to develop a rice cell culture system with high proliferation and screening resistant cell lines of rice to bacterial blight disease caused by Xanthomonas oryza pv. oryzae (Xoo). The culture cells obtained from the callus, cultured on CS-1 medium containing 3 % sucrose and 2 mg L-1 2,4-D for 4 weeks. The results showed that proliferation cell was signifcantly increased 1-fold in 3 weeks of primary culture in CS-1 conditioned medium (fresh/spent medium ratio 1:1) containing 3 % sucrose, 0.5 % glucose, 0.05 % fructose and 2 mg L-1 2,4-D. This medium was used to screen the cell lines through applying culture filtrate of Xoo. The method was to find a novel cell line which could produce high amounts of reactive oxygen species (ROS). Screening results showed 33 % cell lines were strong ROS-producing, two cell lines were selected and cultured for second round screening. The ratio of strong ROS-producing cell lines was increased up to 67 % in the third round screening. The strong ROS-producing cell lines in third round screening can be further cultured for plant regeneration. The rice cell lines with high ROS production may have potential of resistant cell lines against Xoo.

Exploration and Antifungal Assay of Endophytic Fungi as Biocontrol of Onion Purple Blotch Disease Caused by Alternaria porri (Ell) Cif In Vitro Wita Firdausi; Liliek Sulistyowati; Luqman Qurata Aini
AGRIVITA, Journal of Agricultural Science Vol 43, No 1 (2021)
Publisher : Faculty of Agriculture University of Brawijaya in collaboration with PERAGI

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.17503/agrivita.v43i1.2838

Purple blotch disease caused by Alternaria porri is the main destructive foliar disease of genus Allium, causing significant losses in yield of the crops. Recently, purple blotch disease is controlled by synthetic fungicides. However, fungicides have negative effects on the environment. Endophytic fungi can be used as an alternative control because a close symbiosis with the internal tissue of the host can minimize competition in new and complex ecosystems. This study aimed to explore and identify endophytic fungi that have the highest inhibition ability against A. porri and investigate the antagonistic mechanism. The method used in this study is an exploration of endophytic fungi, isolation of A. porri, in vitro antagonism tests, observation of the antagonistic mechanism, extraction of crude protein, SDS-PAGE, and identification. The antagonistic fungi that had the highest inhibition ability were identified as Penicillium citrinum with an inhibitory of 60.04%. Crude protein extracted from P. citrinum which showed inhibitory activity against A. porri is saturation level of ammonium sulfate 80% with a molecular weight of 40 kDa. This study implies that P. citrinum can inhibit the growth of A. porri through its anti fungi compounds. Further in vivo assays or field trials will need to be conducted in future studies.

Mycoparasitic Activity of Indigenous Trichoderma virens Strains Against Mungbean Soil Borne Pathogen Rhizoctonia solani: Hyperparasite and Hydrolytic Enzyme Production Alfi Inayati; Liliek Sulistyowati; Luqman Qurata Aini; Eriyanto Yusnawan
AGRIVITA, Journal of Agricultural Science Vol 42, No 2 (2020)
Publisher : Faculty of Agriculture University of Brawijaya in collaboration with PERAGI

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.17503/agrivita.v0i0.2514

Rhizoctonia solani is one of the harmful pathogens on mungbean, which is very challenging to be controlled. T. virens has been studied intensively and has great potency to control R. solani through mycoparasitism. Seven strains of T. virens isolated from various rhizospheres were tested for their mycoparasitic potential by observing their hyperparasitism and the production of hydrolytic enzymes. All strains showed the ability to suppress the growth of R. solani on dual culture assay as well as on culture filtrate test with the inhibition ability from 43.8 to 68.6% on the dual culture assay and from 22.2 to 71.1% on the culture filtrate assay. Inter-fungal interaction, which was observed by an electron microscope, showed hyperparasitic action of T. virens against R. solani involved the formation of the knob-like structure followed by the growth of Trichoderma hyphae inside host mycelia, coiling, lysed cell wall, and swollen of mycelial tips. Mycoparasitism of T. virens also correlated with the synthesis of hydrolytic enzymes, such as cellulase and chitinase, which influenced the overall hyperparasitic ability of T. virens against the pathogen. Based on in vitro assay, the Tv3 strain proposed as a promising strain to control R. solani due to its high growth inhibition and relatively high cellulase and chitinase productionse.

TRANSFER GEN   -1,3-GLUCANASE DARI JAMUR Trichoderma asperillum PADA KALUS ABAKA UNTUK KETAHANAN TERHADAP PENYAKIT LAYU FUSARIUM RULLY DYAH PURWATI; LILIEK SULISTYOWATI
Jurnal Penelitian Tanaman Industri Vol 18, No 1 (2012): Maret 2012
Publisher : Pusat Penelitian dan Pengembangan Perkebunan

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.21082/jlittri.v18n1.2012.24-30

ABSTRAKKendala utama dalam budidaya tanaman abaka (Musa textilis Nee.)adalah penyakit layu Fusarium yang disebabkan oleh Fusarium oxysporumf.sp cubense (Foc). Upaya perbaikan sifat ketahanan tanaman abakamelalui persilangan sulit dilakukan karena keragaman genetiknya sempitakibat pola perbanyakan secara vegetatif yang terus-menerus.Transformasi gen ketahanan β-1,3-Glucanase merupakan salah satualternatif untuk memperbaiki sifat ketahanan tanaman dengan bantuanvektor Agrobacterium tumefaciens. Gen  -1,3-Glucanase diisolasi darijamur endofit Trichoderma asperillum yang diketahui antagonis terhadapFusarium oxisporum. Penelitian bertujuan untuk mengintroduksi gen β-1,3 Glucanase pada tanaman abaka, sebagai tahap awal untuk memperolehtanaman abaka tahan terhadap penyakit layu Fusarium. Penelitiandilaksanakan di Laboratorium Bioteknologi Fakultas Pertanian, Univer-sitas Brawijaya dan Laboratorium Kultur Jaringan Balai PenelitianTanaman Tembakau dan Serat, mulai Juni 2007 sampai dengan Mei 2009.Penelitian terdiri atas tiga tahap sebagai berikut: transfer gen  -1,3-Glucanase pada kalus abaka embriogenik, regenerasi tunas dan planletabaka transforman, dan konfirmasi planlet abaka transforman yangmengandung gen Gus dan  -1,3-Glucanase. Transfer gen dilakukanmelalui vektor A. tumefaciens strain LBA4404 yang mengandung plasmidpB2GW7 berisi gen-gen  -1,3-Glucanase, Gus (  -glucuronidase) sebagaigen pelapor dan Bar (Basta resistance) sebagai gen penyeleksi. Kalusabaka klon UB-13 embriogenik berukuran 3 x 3 x 3 mm 3 direndam dalamsuspensi A. tumefaciens, kemudian ditanam pada media kokultivasi selama2 hari. Setelah kokultivasi, kalus dipindahkan ke media MS cair+Timentin100 ppm selama 2 minggu. Selanjutnya kalus dipindahkan ke mediainduksi kalus (MK) yaitu MS + BAP 5 mg/l + Thidiazuron 0,4 mg/l +vitamin C 100 mg/l + Basta 50 ppm + Timentin 100 ppm. Regenerasitunas dilakukan dengan memindahkan kalus transforman ke media induksitunas (MT): MS+BAP 0,5 mg/l + vitamin C 100 mg/l dengan penambahandan tanpa Timentin 100 ppm. Tunas transforman dengan tinggi 2-3 cmdipindahkan ke dalam media induksi akar (MA) : MS + arang aktif 2 g/ldengan penambahan dan tanpa Timentin 50 ppm. Keberadaan gen Gusdideteksi dengan reaksi histokimia, dan konfirmasi keberadaan gen  -1,3-Glucanase dilakukan dengan Polymerase Chain Reaction (PCR). Daripenelitian berhasil diperoleh 2% kalus transforman yang lolos seleksiBasta. Hasil konfirmasi keberadaan gen Gus pada planlet transformanmenunjukkan 9 dari 20 (45%) planlet yang diuji, positif mengandung genGus. Konfirmasi keberadaan gen  -1,3-Glucanase dengan PCRmenunjukkan hanya 2 dari 20 planlet transforman, positif mengandung  -1,3-Glucanase. Pengujian ketahanan dari plantlet transgenik tersebut perludilakukan terhadap Fusarium oxisporum f.sp cubense (Foc).Kata kunci: Musa textilis Nee., transformasi gen,  -1,3-Glucanase,Agrobacterium tumefaciens, penyakit, jamur patogen,FusariumABSTRACTThe main constraint of abaca (Musa textilis Nee.) cultivation isinfection of wilt disease caused by Fusarium oxysporum f.sp cubense(Foc). The effort to improve abaca resistance through hybridization is stilldifficult due to narrow genetic variability resulted from continuousvegetative multiplication. Transformation of  -1,3-Glucanase resistancegene is an alternative way to improve character of genetic resistance withhelp of Agrobacterium oxisporum. The research aimed at introducing  -1,3-Glucanase gene to abaca plants prior to obtaining the plants resistanceagainst Fusarium wilt diseases. The research was conducted inBiotechnology Laboratory, Faculty of Agriculture Brawijaya Universityand Tissue Culture Laboratory of Indonesian Tobacco and Fibre CropsResearch Institute, from June 2007 to May 2009. This experimentconsisted of three steps, namely:  -1,3-Glucanase gene transfer onto abacaembriogenic calli, regeneration of transgene abaca shoots and plantlets,and confirmation of transgene abaca plantlets containing Gus and  -1,3-Glucanase genes. Gene transfer was performed using A. tumefaciensvector strain LBA4404 with pB2GW7 containing genes of  -1,3-Glucanase and Gus (  -glucuronidase) as reporter, and Bar (Bastaresistance) as selector marker. Embriogenic calli of abaca clone UB-13were soaked in A. tumefaciens suspension and then cultured in co-cultivation medium for two days. After co-cultivation, calli weretransferred to liquid of MS medium + 100 ppm Timentine for two weeks.Furthermore, the calli were sub-cultured into callus induction medium :MS + 5 mg BAP/l + 0.4 mg Thidiazuron/l + 100 mg vitamin C/l + 50 ppmBasta + 100 ppm Timentine. Shoots regeneration was conducted bytransferring transgene calli to shoot induction medium : MS + 0.5 mg/lBAP + 100 mg vitamin C/l with and without addition of 100 ppmTimentine. Transgene shoots with 2-3 cm height were sub-cultured to rootinduction medium : MS + 2 g active charcoal/l with and without additionof 50 ppm Timentine. Detection of Gus gene was conducted usinghistochemical reaction, while confirmation of  -1,3-Glucanase gene wasperformed by PCR. This project resulted in 2% transgene calli passingBasta selection. Nine out of 20 plantlets (45%) confirmed the existance ofGus gene. PCR results showed that only 2 out of 20 transformed plantlets positively contained  -1,3-Glucanase gene. The plantlets resistanceagainst Fusarium oxisporum f.sp cubense (Foc) needs to be evaluated.Key words: Musa textilis Nee, gene transformation,  -1,3-Glucanase,Agrobacterium tumefaciens, plant disease, fungal disease,Fusarium

IMMUNOGENESITY OF SPESIFIC PROTEIN MOLECULAR WEIGHT 16 KDa (PS16) LEAF OF SIAM CITRUS INFECTED BY CITRUS VEIN PHLOEM DEGENERATION (CVPD) DISEASE Made Sritamin; Aulanni ‘am Aulanni ‘am; IGP. Wirawan; Liliek Sulistyowati
JURNAL PENELITIAN BIOLOGI BERKALA PENELITIAN HAYATI Vol 13 No 1 (2007): December 2007
Publisher : The East Java Biological Society

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.23869/325

Citrus Vein Phloem degeneration (CVPD) is an important citrus disese, which damaged citrus plantation and causing decrease of citrus production. In Indonesia, the CVPD disease caused by Liberobacter asiaticum bactery and the disease spread out by vectir insect Diaphorina citri and using infected bud in wood grafting. In infected citrus plant, two specific protein molecules with molecular weigt 16 kDa and 66 kDa are found. These protein molecules are not found in healthy citrus plant. The immunogenicity of PS16 accumulated on leaf of citrus plant infected by CVPD is known yet. The research material were leaves of citrus plant infected CVPD, leaves of healthy citrus plant and reagent used these research are for isolation of the total protein leaf of citrus plant, SDS-PAGE electroforesis, electroelution of PS16, ELISA Methods, Dot-Blot Method, anti-PS16 as aprimery antibody and secondary antibody is anti-Rabbit IgG Conjugated AP. The result of the research showed that of PS16 accumulated on leaf of citrus plant infected CVPD has immunogenic character. It is indicated by increase of the titer anti-PS16 after first immunization ang 2nd booster by indirect ELISA method and can be used to induce antibody (anti-PS16) and so showed that positive reaction between PS16 with anti-PS16. It is indicated by purples dark blue on cellulose membrane by Dot Blot method.

Purification and Identification of an Antifungal Protein from an Isolated Fungus with Antagonism to Colletotrichum gloeosporioides MC9 Yohana Avelia Sandy; Yo-Chia Chen; Liliek Sulistyowati
AGRIVITA, Journal of Agricultural Science Vol 44, No 2 (2022)
Publisher : Faculty of Agriculture University of Brawijaya in collaboration with PERAGI

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.17503/agrivita.v44i2.2966

Colletotrichum gloeosporioides is the cause of anthracnose disease on mango. This disease becomes more damaging because it economically affects the harvested fruit during the postharvest season. In this research, eight isolates are isolated from the soil of a mango plantation. One of the isolates shows antifungal activity against C. gloeosporioides MC9. This isolate is identified as Penicillium citrinum isolate S1 based on the phylogenetic analysis of ribosomal rRNA sequence. From the culture of this isolate, extracellular filtrates are collected and evaluated for their antifungal activity. The mycelial growth of C. gloeosporioides is significantly inhibited by the culture supernatant of P. citrinum isolate S1. The culture filtrate is used to purify the antifungal protein using ammonium sulfate and ultrafiltration methods. Results show that the antifungal protein was estimated at around 40 kDa molecular weight when separated on a 10% Sodium dodecyl sulfate-polyacrylamide gel. After nine days of incubation, this antifungal protein’s inhibition effect with a concentration of 0.94 mg/ml remained 63.6% against C. gloeosporioides. The LCMS result showed that the antifungal protein belongs to the L-asparaginase superfamily. Based on this result, the antifungal protein produced by P. citrinum S1 has the potential to control mango anthracnose disease caused by C. gloeosporioides.

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