Amplifying and Sequencing Analysis the Internal Transcribed Spacer (ITS) Regions of Olpidium Viciae Kusano’s Ribosomal DNA in Broad Bean

Abstract:

Plasmodiophora fire of broad bean is responsible for Olpidium Viciae Kusano, which is a kind of Fungi subdivided into bacteria flagellum amon. We have developed a polymerase chain reaction based method for the rapid identification internal transcribed spacer (ITS) regions of productionally significant fungi Olpidium Viciae Kusano from areas of 2500~3000 metres above sea level. Sequences of the nuclear internal transcribed spacer (ITS) regions ITS1 and ITS4 have been used widely in molecular characteristic studies because of their relatively high variability and facility of amplification. A universal quickly SDS micro-DNA extraction method was used combining a RNaseA pretreatment step to remove PCR interferential RNA. Target sequences in ITS regions genomic were amplified by PCR and sequenced. Using Hanpanchun lesion and healthy bean leaves as template and ITS1, ITS4 as primer to amplify ITS region, the results revealed ITS gene of broad bean genome could be amplified with size of 750bp from healthy leaves, it could be amplified two fragments of 750bp and 500bp from the DNA template extracted from Hanpanchun lesion tissue. The ITS sequence of Olpidium Viciae Kusano is 99% homoeology with Cercospora (grey speck) pathogen. This may lay the foundation for research about classification and analyze evolutionary relationships of Olpidium Viciae Kusano.

• Periodical: Advanced Materials Research (Volumes 271-273)
• Pages: 507-513
• DOI: https://doi.org/10.4028/www.scientific.net/AMR.271-273.507
• Online since: July 2011
• Authors: Ji Ming Yan, Xiao Hong Shi, Miao Mei, Hong Bo Dai, Hua Zhi Ye
• Keywords: Amplification, Broad Bean, Internal Transcribed Spacer Region (ITS), Olpidium Viciae Kusano, Ribosomal DNA, Sequence Analysis

References

[1] Campbell RN, Sim ST. Host specificity and nomenclature of Olpidium bornovanus (¼Olpidium radicale) and comparisons to Olpidium brassicae. Canadian Journal of Botany Vol. 72 (1994), p.1136–1143.

DOI: 10.1139/b94-139

[2] Lin Dawu, Cui Guangcheng. Suvey on Olpidium viciae Kusamo in Vicia faba in Tibet. Southwest China Journal of Agricultural Sciences, 2(2) (1989).

[3] Ward, E. Use of the polymerase chain reaction for identifying plant pathogens. In Ecology of Plant Pathogens (ed. J. P. Blakeman & B. Williamson), pp.143-160. C.A.B.I. : Wallingford, U. K(1994).

[4] Kularatne HAGC, Lawrie AC, Barber PA, Keane PJ. A specific primer PCR and RFLP assay for the rapid detection and differentiation in planta of some Mycosphaerella species associated with foliar diseases of Eucalyptus globulus. Mycological Research Vol. 108(2004).

DOI: 10.1017/s0953756204000759

[5] E. Ward, M.J. Adams. Analysis of ribosomal DNA sequences of Polymyxa species and related fungi and the development of genus- and species-specific PCR primers. Mycological Research, Vol. 102(1998), pp.965-974.

DOI: 10.1017/s0953756297005881

[6] Pramateftaki, P. V., Lanaridis, P., and Typas, M. A. Molecular identification of wine yeasts at species or strain level: a case study with strains from two vine-growing areas of Greece. J. Appl. Microbiol., Vol. 89(2000), p.236–248.

DOI: 10.1046/j.1365-2672.2000.01102.x

[7] Guillamónd, J.M., Sabaté, J., Barrio, E., Cano, J., and Querol, A. Rapid identification of wine yeast species based on RFLP analysis of the ribosomal internal transcribed spacer (ITS) region. Arch. Microbiol., Vol. 169(1998), p.387–392.

DOI: 10.1007/s002030050587

[8] Gurtler, V., Stanisich, V.A. Newapproaches to typing and identification of bacteria using the 16S-23S rDNA spacer region. Microbiology Vol. 142(1996), p.3–16.

DOI: 10.1099/13500872-142-1-3

[9] Conrads, G., Claros M.C., Citron, D.M., Tyrrell, K.L., Merriam, V., Goldstein, E.J. 16S-23S rDNA internal transcribed spacer sequences for analysis of the phylogenetic relationships among species of the genus Fusobacterium. Int. J. Syst. Evol. Microbiol. Vol. 52(2002).

DOI: 10.1099/00207713-52-2-493

[10] Guasp, C., Moore, E.R., Lalucat, J., Bennasar, A. Utility of internally transcribed 16S-23S rDNA spacer regions for the definition of Pseudomonas stutzeri genomovars and other Pseudomonas species. Int. J. Syst. Evol. Microbiol. Vol. 50(2000).

DOI: 10.1099/00207713-50-4-1629

[11] Motoyama, Y., Ogata, T. 16S-23S rDNA spacer of Pectinatus, Selenomonas and Zymophilus reveal new phylogenetic relationships between these genera. Int. J. Syst. Evol. Microbiol. Vol. 50(2000), p.883–886.

DOI: 10.1099/00207713-50-2-883

[12] Yoon, J.H., Lee, S.T., Kim, S.B., Goodfellow, M., Park, Y.H. Inter- and intraspecific genetic analysis of the genus Saccharomonospora with 16S to 23S ribosomal DNA (rDNA) and 23S to 5S rDNA internally transcribed spacer sequences. Int. J. Syst. Bacteriol. Vol. 47(1997).

DOI: 10.1099/00207713-47-3-661

[13] White TJ, Bruns T, Lee S, Taylor J. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Sninsky JJ, White TJ (eds), PCR Protocols: a guide to methods and amplifications. Academic Press, San Diego(1990).

DOI: 10.1016/b978-0-12-372180-8.50042-1

[14] Lee SB, Taylor JW. Phylogeny of five fungus-like protoctistan phytophthora species, inferred from the internal transcribed spacer of ribosomal DNA. Molecular Biology and Evolution Vol. 9(1992) p.636–653.

DOI: 10.1093/oxfordjournals.molbev.a040750

[15] Sreenivasaprasad S, Mills PR, Meehan BM, Brown AE, 1996. Phylogeny and systematics of 18 Colletotrichum species based on ribosomal DNA spacer sequences. Genome Vol. 39(1996), p.499–512.

DOI: 10.1139/g96-064

[16] Crous PW, Hong L, Wingfield BD, Wingfield MJ. ITS rDNA phylogeny of selected Mycosphaerella species and their anamorphs occurring on Myrtaceae. Mycological Research Vol. 105(2001) p.425–431.

DOI: 10.1017/s0953756201003835

[17] Dunne CP, Glen M, Tommerup IC, Shearer BL, Hardy GESt J. Sequence variation in the rDNA ITS of Australian Armillaria species and intra-specific variation in A. luteobubalina. Austra. Plant Pathol. Vol. 31(2002) p.241–251.

DOI: 10.1071/ap02015

[18] Boyang Cao, Min Wang, Lei Liu, Zhemin Zhou, Shaoping Wen, Antoni Rozalski, Lei Wang. 16S-23S rDNA internal transcribed spacer regions in four Proteus species. Journal of Microbiol. Methods. Vol. 77(2009), pp.109-118.

DOI: 10.1016/j.mimet.2009.01.024

[19] Wang Lihua, Yin Fuyou, Liu Jimei, Ye Lǚrong. Genetic diversity and relationships of the wild buckwheat resources from yunnan province revealed by RAPD. Buckwheat News, Vol. (2) (2004), pp.7-15.

[20] Wang Jianbo, Zhang Wenju, Cheng Jiakuan. Application of ITS sequences of nuclear rDNA in phylogenetic and evolutionary studies of angiosperms. Acta Phytotaxonomica Sinica, Vol. 37(4)(1999), p.407.

[21] Somai BM, Dean RA, Farnham MW, Zitter TA, Keinath AP. Internal transcribed spacer regions 1 and 2 and random amplified polymorphic DNA analysis of Didymella bryoniae and related Phoma species isolated from curcubits. Phytopathology, Vol. 92(2002).

DOI: 10.1094/phyto.2002.92.9.997

[22] Zambino PL, Szabo LJ. Phylogenetic relationships of selected cereal and grass rusts based on rDNA sequence analysis. Mycologia, Vol. 85(1993), p.401.

DOI: 10.2307/3760702

[23] Steven VV, Stephen RP. Testing the utility of internally transcribed spacer sequences in coral phylogenetics. Molecular Ecol, Vol. 13(2004), p.2763.

DOI: 10.1111/j.1365-294x.2004.02265.x

[24] Liu Zhong, Wang Xiaoquan, Chen Zhiduan. The Phylogeny of Schisandraceae Inferred from Sequence Analysis of the nrDNA ITS Region. Acta Botanica Sinica, Vol. 42(7)(2000), p.758.

[25] Gardes MT, Bruns D. ITS primers with enhanced specificity for basidiomycetes application to the identification of mycorrhizae and rusts. Molecular Ecol, Vol. 2(1993), p.113.

DOI: 10.1111/j.1365-294x.1993.tb00005.x

[26] Zhao Jie. ITS sequential analysis and its application in molecular detection of plant fungal disease. Shaanxi Journal of Agricultural Sciences, Vol. (4)(2004), p.35.

[27] Cai Jinna, Zhou Kaiya, Xu Luoshan, Wang Zhengtao, Shen Xi, Wang Yiquan, Li Xiaobo. Ribosomal DNA its sequence analyses of Cnidium monnieri from different geographical origin in China. Acta Pharmaceutica Sinica, Vol. 35(1)(2000), p.56.

[28] Chinese Academy of Sciences-Flora Repubulicae Popularis Sinicae editorial board, Flora Repubulicae Popularis Sinicae(30-1). Beijing.

DOI: 10.1051/978-2-7598-2503-5.c009

[29] Lucy M. H., Paul J. H. and John A. W. A comparison of Olpidium isolates from a range of host plants using internal transcribed spacer sequence analysis and host range studies. Fungal Biology, Vol. 114(2010), pp.26-33.

DOI: 10.1016/j.mycres.2009.09.008

[30] Jung T, Nechwatal J, Cooke DEL, Hartmann G, Blaschke M, OsswaldWF, Duncan JM, Delatour C. Phytophthora pseudosyringae sp nov., a new species causing root and collar rot of deciduous tree species in Europe. Mycological Research Vol. 107(2003).

DOI: 10.1017/s0953756203008074

[31] Muthumeenakshi S, Goldstein AL, Stewart A, Whipps JM. Molecular studies on intraspecific diversity and phylogenetic position of Coniothyrium minitans. Mycological Research Vol. 105(2001), p.1065–1074.

DOI: 10.1016/s0953-7562(08)61968-1