تشخیص مولکولی ژن‌های القاء شده گیاه سیب زمینی دخیل در دفاع علیه باکتری Ralstonia solanacearum

نوع مقاله : مقاله کامل پژوهشی

نویسندگان

1 مؤسسه تحقیقات ثبت و گواهی بذر و نهال، سازمان تحقیقات، آموزش و ترویج کشاورزی، کرج، ایران.

2 مؤسسه تحقیقات اصلاح و تهیه نهال و بذر، سازمان تحقیقات، آموزش و ترویج کشاورزی، کرج، ایران.

3 گروه بیماری‌شناسی گیاهی، دانشکده کشاورزی، دانشگاه تربیت مدرس، تهران، ایران.

چکیده

با استفاده از تکنیک cDNA-AFLP واکنش مولکولی رقم حساس (مارفونا) و رقم متحمل (الس) در گیاه سیب زمینی (Solanum tuberosum) و ژنوتیپ مقاوم (S. phureja) برای شناسایی ژن­های دخیل در مکانیسم­های دفاعی مورد بررسی قرار گرفت. بیان ژن­های مرتبط با دفاع در چهار زمانهای 0، 2، 4 و 7 روز بعد از آلوده سازی ریشه گیاهان با باکتری R. solanacearum در شرایط درون شیشه­ای بررسی گردید. حدود 2820 قطعه مشتق از رونوشت مشاهده شد که در این میان 206 قطعه الگوهای بیان متفاوتی نشان دادند. در مجموع 20 قطعه با بیشترین اختلاف، توالی­یابی شدند و همولوژی آنها با ژن­های شناخته شده قبلی در بانک­ ژن بررسی گردید. این ژن­ها تشابه قابل توجهی با برخی ژن­های درگیر در فرایند­های فیزیولوژیکی واکنش به استرس­ها، دفاع، ژن‌های ساختاری وژن‌های دخیل در نسخه­برداری، سنتز و بسته­بندی پروتئین داشتند. تغییر بیان این ژن­ها در S. phureja نسبت به  S. tuberosum پس از آلودگی با R. solanacearum بیشتر بود.

کلیدواژه‌ها


عنوان مقاله [English]

Molecular characterization of defense-related genes, induced against Ralstonia solanacearum in potato

نویسندگان [English]

  • C. Moslemkhani 1
  • J. Mozafari 2
  • M. Shams Bakhsh 3
  • E. Mohamadi Goltape 3
1 Seed and Plant Certification and Registration Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
2 Seed and Plant Improvement Institute, Agricultural Research, Education and Extension Organization (AREEO), Ka-raj, Iran.
3 Department of Plant Pathology, Faculty of Agriculture, Tarbiat Modares University, Tehran, Iran.
چکیده [English]

Molecular responses of susceptible (Marfona) and tolerant (Els) cultivars of Solanum tuberosum, and a resistant genotype of S. phureja were comparatively studied using a cDNA-AFLP technique to identify candidate genes involved in defense mechanisms against Ralstonia solanacearum. Transcriptional changes were studied at 0, 2, 4 and 7 days post inoculation of the plants by R. solanacearum through root inoculation under in vitro conditions. Of the 2820 transcript derived fragments (TDFs) detected, 206 TDFs showed variable patterns. In total 20 TDFs with stronger differential induction were sequenced. These TDFs showed significant homologies with the genes involved in physiological processes, stress responses, cell structure, defense, transcription and protein synthesis/folding. Approximately 5% of the sequenced TDFs were identified with unknown function. Induction of a higher number of genes was altered in response to R. solanacearum in S. phureja compared to S. tuberosum.

کلیدواژه‌ها [English]

  • Bacterial wilt
  • Solanum tuberosum
  • S. phureja
  • cDNA-AFLP
Allen C., Prior P. and Hayward A. C. 2005. Bacterial wilt disease and the Ralstonia solanacearum species
complex. St. Paul, MN: APS Press.
Ausubel F. M., Brent R., Kingston R. E., Moore D. D., Seidman J. G., Smith J. A. and Struhl K. 1992. Current protocols in molecular biology. New York: Greene Publishing Association; Wiley-Interscience, v.1. Bachem C. W. B., Oomen R. J. F. and Visser R. 1998. Transcript imaging with cDNA-AFLP: A step by step protocol.  Plant Molecular Biology Reporter 16: 157- 173.
Banzet N., Richaud C. Deveaux Y. Kazmaier M. Gagnon J., and Triantaphylides C. 1998. Accumulation of small heat shock proteins, including mitochondrial HSP22, induced by oxidative stress and adaptive response in tomato cells. Plant Journal 13:519–527.
Bassam B. J., Caetano-Anolles G. and Gresshoff P. M. 1991. Fast and sensitive silver staining of DNA in polyacrylamide gels. Analytical Biochemistry 196:80-83.
Boshou L. 2005. A broad review and perspective on breeding for resistance to bacterial wilt. In Bacterial Wilt Disease and the Ralstonia solanacearum SpeciesComplex, eds. Allen, C., Prior, P. and Hayward, A. C., American Phytopathological Society, St. Paul, MN, USA, 225-238.
Byth H. A. Kuun K. G. and Bornma N. L. 2001. Virulence-dependent induction of Hsp70/Hsc70 in tomato by Ralstonia solanacearum . Plant Physiology and Biochemistry 39:697–705
Chester K. S. 1993. The problem of acquired physiological immunity in plants. The Quarterly Review of Biology 8:129-154.
Choudhary M. K. Basu D., Datta A., Chakraborty N. and Chakraborty, S., 2009. Dehydration-responsive nuclear proteome of rice (Oryza sativa L.) illustrates protein network, novel regulators of cellular adaptation and evolutionary perspective. Molecular and Cellular Proteomics 8:1579–1598.
Dahal D., Heinz D., Dorsselaer A. V., Braun H. P. and Wydra, K. 2009. Pathogenesis and stress related, as well as metabolic proteins are regulated in tomato stems infected with Ralstonia solanacearum. Plant Physiology and Biochemistry 47:838-846.
Dahal D., Pich A., Braun H. P. and Wydra, K., 2010. Analysis of cell wall proteins regulated in stem of susceptible and resistant tomato species after inoculation with Ralstonia solanacearum, a proteomic approach. Plant Molecular Biology 73:643-658.
Denny T. P., 2006. Plant Pathogenic Ralstonia species. Plant Associated Bacteria, (573-644). APS Press.
Esposito N., Ovchinnikova O. G., Barone A., Zoina A., Holst O. and Evidente, A. 2008. Host and non-host plant response to Bacterial wilt in potato: role of the lipopolysaccharide isolated from Ralstonia solanacearum and molecular analysis of plant pathogen interaction. Chemistry and Biodiversity 5:2662-2675.
Fock I., Collonnier C., Purwito A.,  Luisetti J., Souvannavong V., Vedel F., Servaes A., Ambroise A., Kodja H., Ducreux G. and Sihachakr, D. 2000. Resistance to bacterial wilt in somatic hybrids between Solanum tuberosum and Solanum phureja. Plant Science 160:165-175.
Fock I., Luisetti J., Collonnier C., Vedel F., Ducreux G., Kodja H. and Sihachakr D. 2005. Solanum phureja and S. stenotomum are sources of resistance to Ralstonia solanacearum for somatic hybrids of potato. Page 253-259 in Bacterial wilt disease and the Ralstonia solanacearum species complex. C. Allen, P. Prior, and A.C. Hay ward (eds.), St. Paul, MN, USA; Am Phytophatology Soc. (APS Press).
Guevara M. G., Verıssimo P., Pires E., Faro C. and Daleo G. R. 2004. Potato aspartic proteases: induction, antimicrobial activity and substrate specificity. The Journal of Plant Pathology 86: 233–238.
Kiba A., Tomiyama H., Takashi H., Ohnishi K., Okuno T. and Hikichi Y. 2003. Induction of resistance and expression of defense-related genes in tobacco leaves infiltrated with Ralstonia solanacearum. Plant and Cell Physiology 44: 287–295
Kolomiets M. V., Chen H., Gladon R. J., Braun E. J. and Hannapel D. J., 2000. A leaf lipoxygenase of potato induced specifically by pathogen infection. Plant Physiology 124:1121–1130.
Li G. C., Jin L. P., Wang X. W., Xie K. Y., Yang Y., Van der Vossen A. G., Huang S. W. and Qu D. Y. 2010. Gene transcription analysis during interaction between potato and Ralstonia solanacearum. Russian Journal of Plant Physiology 57:685-695.
Li G. C., Jin L. P., Wang X. W., Xie K. Y., Xie B. Y. and Qu D. Y.  2007. Cloning of DnaJ-like gene cDNA in diploid potato using RACE methods combined with cDNA library. Acta Horticulturae Sinica 34: 649-654
Matteo A. D., Federici L., Mattei B., Salvi G., Johnson A. K., Savino C.,  Lorenzo G. D. and Tsernoglou D. 2003. The crystal structure of polygalacturonase-inhibiting protein (PGIP), a leucine-rich repeat protein involved in plant defense. Proceedings of the National Academy of Sciences 100:10124–10128.
Moslemkhani C., Mozafari J. Alizadeh, A. 2005. Diagnosis of Ralstonia solanacearum in potato seed tubers and soil, using PCR technique. Iranian Journal of Plant Pathology 47:215-228.
Moslemkhani C., Mozafari J., Alizadeh A., Shamsbakhsh M. and Mohamadi Goltapeh E. 2011. Evaluation of potato cultivars for resistance to Ralstonia solanacearum under in vitro and greenhouse conditions. Seed and Plant Improvement Journal 26:501-515.
Moslemkhani K., Mozafari J., Shams Bakhsh M. and Mohamadi Goltapeh E. 2012. Expressions of some defense genes against Ralstonia solanacearum in susceptible and resistant potato genotypes under in vitro conditions. Iranian Journal of Plant Pathology 4:183-197.
Moslemkhani K. and Mozafari J. 2014. Effects of various parameters influencing the quality of results in a cDNA-AFLP technique used for transcriptom analysis of potato. Journal of agricultural biotechnology 6: 183-197
Murashige T. and Skoog F. 1962. A revised medium for rapid growth and bio assay with tobacco tissue culture. Physiological Planta 15: 473-479.
Nie X. and Singh R. P., 2001. Differential accumulation of Potato Virus A and expression pathogenesis related genes in resistant potato cv. Shepody upon graft inoculation. Phytopathology 91:197-203.
Riechmann J. L., Heard J., Martin G., Reuber L., Jiang C., Keddie J., Adam L., Pineda O., Ratcliffe O. J. and Samaha R. R. 2000. Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes. Science 29:2105-2110
Rowe P. R., Sequeira L. and Gonzalez L. C. 1972. Additional genes for resistance to Pseudomonas solanacearum in Solanum phureja. Phytopathology 62:1093-1094.
Rushton P. J. and Somssich I. E. 1998 Transcriptional control of plant genes responsive to pathogens. Current Opinion in Plant Biology 1:311-315.
Schaad N. W., Jones J. and Band Chun, W. 2001. Laboratory guide for identification of plant pathogenic bacteria (3rd ed.). The American Phytopathological Society, St. Paul, Minn: APS.
Schacht T., Unger C., Pich A. And Wydra K., 2011. Endo and exopolygalacturonases of Ralstonia solanacearum are inhibited by polygalacturonase inhibiting protein activity in tomato stem extracts. Plant Physiology et biochemistry. Plant Physiology and Biochemistry 49:377 -387
Virupaksh U., Patil J., Gopal J. and Singh B.P. 2012. Improvement for bacterial wilt resistance in potato by conventional and biotechnological approaches. Agricultural research 1:299-316.
Vos P., Hogers R. and Bleeker M. 1995. AFLP: a new technique for DNA fingerprinting. Nucleic Acid Research 23:4407-4414.
Zuluaga A. P., Sole M., Lu H., Gongora-Castillo E., Vaillancourt B., Coll N., Buell C. B. and Valls M. 2015. Transcriptome responses to Ralstonia solanacearum infection in the roots of the wild potato Solanum commersonii. BMC Genomics 16:246.