Recombinant antibody production by cloning of Pepper yellow leaf curl Indonesia virus (PepYLCIV) coat protein gene

Authors

  • Yashanti Berlinda Paradisa Research Center for Biotechnology, National Research and Innovation Agency
  • Sri Sulandari Department of Plant Protection, Faculty of Agriculture, Universitas Gadjah Mada
  • Sedyo Hartono Department of Plant Protection, Faculty of Agriculture, Universitas Gadjah Mada
  • Susamto Somowiyarjo Department of Plant Protection, Faculty of Agriculture, Universitas Gadjah Mada
  • Mery Windarningsih Faculty of Agriculture, University of Mataram
  • Dini Wahyu Kartika Sari Departement of Fisheries, Faculty of Agriculture, Universitas Gadjah Mada
  • Christina Retna Handayani Main Center for Brackish Water Aquaculture Development Jepara

DOI:

https://doi.org/10.23960/jhptt.1221-13

Keywords:

Begomovirus, chili, polyclonal antibody, recombinant protein

Abstract

Pepper yellow leaf curl Indonesia virus (PepYLCIV) is an important pathogen on chili cultivation and is transmitted through the seed. Serological tests are sensitive, accurate, efficient and it has been widely used for the detection of seed-transmitted plant viruses. This study aimed to produce PepYLCIV recombinant protein as a material to produce recombinant antibodies PepYLCIV. DNA was extracted from infected chili leaves collected from Congkrang, Muntilan, Central Java verified using primer PepYLCIV-BamHI and PepYLCIV-EcoRI and produced an amplicon at 840 bp. The amplified fragments were cloned into the pET32a then transformed to Escherichia coli BL21. The percentage of nucleotide sequence identity and sequence of amino acid, PepYLCIV CK-6 isolates had the highest similarity of nucleotide and amino acid sequences to of chili isolates from Bandung. The expressed recombinant protein was obtained with IPTG concentration 0,5 mM and harvested at 6 hours after IPTG induction. SDS PAGE analysis of the recombinant plasmid Begomovirus CK-6 showed that the coat protein size was about 29 kDa. Immunization was carried out on rabbits by injecting 150 µg of recombinant protein 4 times with an interval of 1 week to produce crude antiserum and pure antiserum capable of detecting PepYLCIV in chili and Ageratum conyzoides using I-ELISA and DIBA tests.

References

Abouzid AM, Freitas-Astua J, Purcifull DE, Polston JE, Beckham KA, Crawford WE, Petersen MA, Peyser B, Patte C, & Hiebert E. 2002. Serological studies using polyclonal antisera prepared against the viral coat protein of four Begomoviruses expressed in Escherichia coli. Plant Dis. 86(10): 1109–1114. https://doi.org/10.1094/PDIS.2002.86.10.1109

Anggraini S & Hidayat S. 2014. Sensitivitas metode serologi dan polymerase chain reaction untuk mendeteksi Bean common mosaic potyvirus pada kacang panjang [Sensitifity of serology and polymerase chain reaction methods for detection of Bean common mosaic potyvirus in yard long bean]. J. Fitopatol. Indones. 10(1): 17–22. https://doi.org/10.14692/jfi.10.1.17

Astuti NT, Darsono N, Widyaningrum S, Sawitri WD, Wahyuningsih SPA, & Darmanto W. 2019. Expression and purification of recombinant coat protein of Sugarcane mosaic virus from Indonesian isolate as an antigen for antibody production. Indones. J. Biotechnol. 24(1): 57–64. https://doi.org/10.22146/ijbiotech.45551

Bartkus J. 2016. DNA sequencing for clinical and public health virology: some assembly required. In: Loeffelholz MJ, Hodinka RL, Young SA, & Pinsky BA (Eds.). Clinical Virology Manual. Fifth Edition. pp. 173–199. ASM Press, Washington, DC.

Boonham N, Kreuze J, Winter S, van der Vlugt R, Bergervoet J, Tomlinson J, & Mumford R. 2014. Methods in virus diagnostics: from ELISA to next generation sequencing. Virus Res. 186: 20–31. https://doi.org/10.1016/j.virusres.2013.12.007

Brandariz-Fontes C, Camacho-Sanchez M, Vilà C, Vega-Pla JL, Rico C, & Leonard JA. 2015. Effect of the enzyme and PCR conditions on the quality of high-throughput DNA sequencing results. Sci. Rep. 5: 8056. https://doi.org/10.1038/srep08056

Brown JK, Fauquet CM, Briddon RW, Zerbini M, Moriones E, & Navas-Castillo J. 2012. Family Geminiviridae. In: King AMQ, Adams MJ, Carstens EB, & Lefkowitz EJ (Eds.). Virus Taxonomy: Ninth Report of the International Committee on Taxonomy of Viruses. pp. 351–373. Elsevier Academic Press, San Diego.

Buckhout-white S, Person C, Medintz IL, & Goldman ER. 2018. Restriction enzymes as a target for DNA-based sensing and structural rearrangement. ACS Omega. 3(1): 495–502. https://doi.org/10.1021/acsomega.7b01333

Dellaporta SL, Wood J, & Hicks JB. 1983. A plant DNA minipreparation: Version II. Plant Mol. Biol. Rep. 1(4): 19–21. https://doi.org/10.1007/BF02712670

Donovan RS, Robinson CW, & Glick BR. 1996. Review: Optimizing inducer and culture conditions for expression of foreign proteins under the control of the lac promoter. J. Ind. Microbiol. 16(3): 145–154. https://doi.org/10.1007/bf01569997

Fadhila C, Lal A, Vo TTB, Ho PT, Hidayat SH, Lee J, Kil EJ, & Lee S. 2020. The threat of seed-transmissible Pepper yellow leaf curl Indonesia virus in chili pepper. Microb. Pathog. 143: 104132. https://doi.org/10.1016/j.micpath.2020.104132

Ganefianti DW, Hidayat SH, & Syukur M. 2017. Susceptible phase of chili pepper due to yellow leaf curl Begomovirus infection. Int. J. Adv. Sci. Eng. Inf. Technol. 7(2): 594–601. http://dx.doi.org/10.18517/ijaseit.7.2.1872

Gaur A, Kumar A, Kiran R, & Kumari P. 2020. Importance of seed-borne diseases of agricultural crops: economic losses and impact on society. In: Kumar R & Gupta A (Eds.). Seed-Borne Diseases of Agricultural Crops: Detection, Diagnosis & Management. pp. 3–24. Springer, Singapore.

Grodzki AC & Berenstein E. 2010. Antibody Purification: Ammonium sulfate fractionation or gel filtration. In: Oliver C & Jamur MC (Eds.). Immunocytochemical Methods and Protocols. Third Edition. pp.15–26. Humana Press, New York.

Guerrero J, Regedanz E, Lu L, Ruan J, Bisaro DM, & Sunter G. 2020. Manipulation of the plant host by the Geminivirus AC2/C2 Protein, a central player in the infection cycle. Front Plant Sci. 11(591): 1–18. https://doi.org/10.3389/fpls.2020.00591

Hamdayanty H, Hidayat SH, & Damayanti TA. 2016. Expression of recombinant Sugarcane streak mosaic virus coat protein gene in Escherichia coli. HAYATI J. Biosci. 23(3): 111–116. https://doi.org/10.1016/j.hjb.2016.11.001

Hibi T & Saito Y. 1985. A Dot immunobinding assay for the detection of Tobacco mosaic virus in infected tissues. J. Gen. Virol. 66: 1191–1194.

Inouce-Nagata AK, Nagata T, De Avila AC, & Giordano LDB. 2007. A reliable begomovirus inoculation method for screening Lycopersicon esculentum lines. Hortic. Bras. 25(3): 447–450. https://doi.org/10.1590/S0102-05362007000300024

Islam W. 2017. Management of plant virus diseases; farmer’s knowledge and our suggestions. Hosts and Viruses. 4(2): 28–33. http://dx.doi.org/10.17582/journal.bjv/2017/4.2.28.33

Jamsari J & Pedri J. 2013. Complete nucleotide sequence of DNA a-like genome and DNA-? of monopartite Pepper yellow leaf curl virus, a dominant Begomovirus infecting Capsicum annuum in West Sumatera Indonesia. Asian J. Plant Pathol. 7: 1–14. https://dx.doi.org/10.3923/ajppaj.2013.1.14

Jamsari, Syukriani L, Utami HP, Herberg F, Nellen W, & Ferita I. 2015. Injection technique could as a new promising method for artificial infection of Geminivirus particles in chili pepper (Capsicum annuum L.). Asian J. Agric. Res. 9: 23–32. https://dx.doi.org/10.3923/ajar.2015.23.32

Jones RAC. 2014. Plant virus ecology and epidemiology: historical perspectives, recent progress and future prospects. Ann. Appl. Biol. 164(3): 320–347. https://doi.org/10.1111/aab.12123

Koeda S, Homma K, Tanaka Y, Onizaki D, Kesumawati E, Zakaria S, & Kanzaki S. 2018. Inoculation of capsicums with Pepper yellow leaf curl Indonesia virus by combining agroinoculation and grafting. Hort. J. 87(3): 364–371. http://dx.doi.org/10.2503/hortj.OKD-137

Koenig R. 1981. Indirect ELISA methods for the broad specificity detection of plant viruses. J. Gen. Virol. 55(1): 53–62. https://doi.org/10.1099/0022-1317-55-1-53

Kumar R, Gupta A, Srivastava S, Devi G, Singh VK, Goswami SK, Gurjar MS, & Aggarwal R. 2020. Diagnosis and Detection of Seed-Borne Fungal Phytopathogens. In: Kumar R & Gupta A (Eds.). Seed-Borne Diseases of Agricultural Crops: Detection, Diagnosis & Management. pp. 107–229. Springer, Singapore. https://doi.org/10.1007/978-981-32-9046-4_5

Kumar S, Udaya Shankar AC, Nayaka SC, Lund OS, & Prakash HS. 2011. Detection of Tobacco mosaic virus and Tomato mosaic virus in pepper and tomato by multiplex RT-PCR. Lett. Appl. Microbiol. 53(3): 359–363. https://doi.org/10.1111/j.1472-765x.2011.03117.x

Laprom A, Nilthong S, & Chukeatirote E. 2019. Incidence of viruses infecting pepper in Thailand. Biomol. Concepts. 10(1): 184–193. https://doi.org/10.1515/bmc-2019-0021

Lestari SM, Hidayat SH, & Widodo. 2018. Determination of endophytic fungi as induce resistance agent of chilli pepper against pepper yellow leaf curl disease. Agrivita. 40(2): 249–256. http://doi.org/10.17503/agrivita.v40i2.989

Li X. 2018. Bioengineering of FGFs and new drug developments. In: Li X (Ed.). Fibroblast Growth Factors. pp. 477–558. Elsevier Inc. Amsterdam, Netherlands. https://doi.org/10.1016/B978-0-12-816142-5.00008-4

Liu Z & Yang PC. 2012. Construction of pET-32 ? (+) vector for protein expression and purification. North. Am. J. Med. Sci. 4(12): 651–655. http://doi.org/10.4103/1947-2714.104318

Malathi VG. 2017. Begomovirus: an introduction. In: Saxena S & Tiwari AK (Eds.). Begomoviruses: Occurrence and Management in Asia and Africa. pp. 3–9. Springer, Singapore. https://doi.org/10.1007/978-981-10-5984-1_1

Morra MR & Petty ITD. 2000. Tissue specificity of Geminivirus infection is genetically determined. Plant Cell. 12(11): 2259–2270. https://doi.org/10.1105/tpc.12.11.2259

Munandar M, Romano, & Usman. 2017. Faktor–faktor yang mempengaruhi permintaan cabai merah di Kabupaten Aceh Besar [Analysys of factors influencing the demand of red chilli in Aceh Besar Regency]. Jurnal Ilmiah Mahasiswa Pertanian. 2(3): 80–91. https://doi.org/10.17969/jimfp.v2i3.3752

Neriya Y, Izumi R, Wilisiani F, Hartono S, Wirya GNAS, Nishigawa H, & Natsuaki T. 2020. Complete genome sequence of a Pepper yellow leaf curl Indonesia virus isolated from tomato in Bali, Indonesia. Microbiol. Resour. Announc. 9(25): e00486-20. https://doi.org/10.1128/MRA.00486-20

Nordenstedt N, Marcenaro D, Chilagane D, Mwaipopo B, Rajamäki ML, Nchimbi-Msolla S, Njau PJR, Mbanzibwa DR, & Valkonen JPT. 2017. Pathogenic seedborne viruses are rare but Phaseolus vulgaris endornaviruses are common in bean varieties grown in Nicaragua and Tanzania. PLoS One. 12(8): e0184263. https://doi.org/10.1371/journal.pone.0184263

Noueiry A, Lucas WJ, & Gilbertson RL. 1994. Two proteins of a plant DNA virus coordinate nuclear and plasmodesmal transport. Cell. 76(5): 925–932. https://doi.org/10.1016/0092-8674(94)90366-2

Pratiwi RD. 2019. Optimasi ekspresi human Epidermal Growth Factor (h-EGF) rekombinan dalam Escherichia coli BL21(DE3) dengan variasi media dan konsentrasi penginduksi [in Indonesian]. Chimica et Natura Acta. 7(2): 91–97. https://doi.org/10.24198/cna.v7.n2.23824

PUSDATIN. 2020. Outlook Cabai. Sekteratiat Jenderal Kementrian Pertanian [in Indonesian]. Website: http://epublikasi.pertanian.go.id/download/file/586-outlook-cabai-besar-2020. Accessed 8 April 2021.

Rosano GL & Ceccarelli EA. 2014. Recombinant protein expression in Escherichia coli: advances and challenges. Front. Microbiol. 5: 172. https://doi.org/10.3389/fmicb.2014.00172

Roshan P, Kulshreshtha A, & Hallan V. 2017. Genome Organization of Begomovirus. In: Saxena S & Tiwari AK (Eds.). Begomoviruses: Occurrence and Management in Asia and Africa. pp. 11–32. Springer, Singapore. https://doi.org/10.1007/978-981-10-5984-1_2

Sambrook J, Fritsch EF, & Maniatis T. 1989. Molecular Cloning: a Laboratory Manual. Second Edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.

Sambrook J & Russell DW. 2001a. Molecular Cloning: a Laboratory Manual Vol 1. Third Edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.

Sambrook J & Russell DW. 2001b. Molecular Cloning: a Laboratory Manual Vol 3. Third Edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.

Sastry KS. 2013. Detection of plant viruses in seeds. In: KS Sastry (Ed.). Seed-Borne Plant Virus Diseases. pp. 101-163. Springer Science & Business Media, India.

Selangga DGW & Listihani L. 2021. Molecular identification of Pepper yellow leaf curl Indonesia virus on chili pepper in Nusa Penida Island. J. HPT Tropika. 21(2): 97–102. https://doi.org/10.23960/jhptt.22197-102

Stapleton S, O’Kennedy R, & Tully E. 2005. Immunoassays: production of antibodies. In: Worsfold P, Townshend A, & Poole C (Eds.). Encyclopedia of Analytical Science. Second Edition. pp. 306-316. Elsevier Ltd. Amsterdam. https://doi.org/10.1016/B0-12-369397-7/00263-6

Stills HF. 2012. Polyclonal antibody production. In: Suckow MA, Steven KA, & Wilson RP (Eds.). The Laboratory Rabbit, Guinea Pig, Hamster, and Other Rodents. pp. 259–274. Elsevier Inc. Oxford. https://doi.org/10.1016/C2009-0-30495-X

Sudiono, Yasin N, Hidayat SH, & Hidayat P. 2005. Penyebaran dan deteksi molekuler virus gemini penyebab penyakit kuning pada tanaman cabai di Sumatera [The distribution and molecular detection of geminivirus pathogen of chilli yellowing disease in Sumatera lsland]. J. HPT Tropika. 5(2): 113–121. https://doi.org/10.23960/j.hptt.25113-121

Sulandari S. 2004. Karakterisasi biologi, serologi dan analisis sidik jari DNA virus penyebab penyakit daun keriting kuning cabai [Biological characterization, scrological assay and DNA finger printing analysis of Pepper yellow leaf curl virus]. Dissertation. Institut Pertanian Bogor. Bogor.

Veniari NK, Yuliadhi KA, Nyana IDN, & Suastika G. 2015. Deteksi Cucumber mosaic virus (CMV) dan Chili veinal mottle virus (ChiVMV) pada gulma Commelina spp. di pertanaman cabai (Capsicum spp.) melalui teknik uji serologi dan molekuler [Detection of Cucumber mosaic virus (CMV ) and Chilli veinal mottle virus (ChiVMV) on weed Commelina spp. in cropping chilli pepper (Capsicum spp.) through serology and molecular test]. Jurnal Agroekoteknologi Tropika. 4(1): 45–52.

Viswanathan R, Karuppaiah R, & Ganesh Kumar V. 2011. Expression of Sugarcane streak mosaic virus (SCSMV) coat protein in expression vector as a fusion protein with maltose binding protein expression vector as a fusion protein with maltose binding protein. J. Sugar. Res. 1(1): 63–68.

Weber J, Peng H & Rader C. 2017. From rabbit antibody repertoires to rabbit monoclonal antibodies. Exp. Mol. Med. 49(3): e305. https://doi.org/10.1038/emm.2017.23

Wyatt SD & Brown JK. 1999. Detection of subgroup III Geminivirus isolates in leaf extracts by degenerate primers and polymerase chain reaction. Phytopathology. 86(12): 1288–1293. https://doi.org/10.1094/Phyto-86-1288

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2022-03-01
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