PCR-DGGE and PCR-RFLP Analyses of the Internal Trascribed Spacer(ITS) of Ribosomal DNA in the Genus Rhizopus

  • Published : 2003.06.01

Abstract

To estimate genetic relationships within the genus Rhizopus, genetic variations in 20 strains were investigated by DGGE and PCR-RFLP of rDNA ITS region (ITSI, ITS2,5.8S). The size of the amplified products showed the interspecific polymorphisms, 650 bp,700 bp, and 900 bp. The DGGE approach allowed the separation of PCR amplicons of the same length according to their sequence variations. When the rDNA ITS region was digested with six restriction enzymes, 20 strains were classified into five RFLP haplotypes. The range of similarity between the 20 strains by PCR-RFLP was 42.3-100%. Based on the results of DGGE aud PCR-RFLP, the 20 strains were divided into four groups, R. oryzae, R. stolonifer, R. microsporus and R. homothallicus. Further study of R. homothallicus is required.

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References

  1. Teor. Appl. Genet. v.89 Association of a DNA marker with Hessian fly resistance gene H9 in wheat Dweika,I.M.;H.Ohm;S.Mackenzie;F.Patterson;S.Cambron;R.Ratcliffe
  2. Mycologia. v.73 Sporangiospore ornamentation of thermophilic Rhizopus species and some allied genera Ellis,D.H. https://doi.org/10.2307/3759604
  3. Mycologia. v.77 Species and varieties in the Rhizopus arrhizus-Rhizopus oryzae group as indicated by their DNA complementarily Ellis,J.J. https://doi.org/10.2307/3793074
  4. Mucorales Hesseltine,C.W.;J.J.Ellis;G.C.Ainsworth(ed.);F.K.Sparrow(ed.);A.S.Sussman(ed.)
  5. J. Gen. Appl. Microbiol. no.Suppl.11 Taxonomical studies on genus Rhizopus Inui,T.;Y.Takeda;H.Iizuka
  6. Appl. Environ. Microbiol. v.63 Detection and characterization of fungal infections of Ammophila arenaria (Marram Grass) roots by denturing gradient gel electrophoresis of specifically amplified 18S rDNA Kowalchuk,G.A.;S.Gerards;J.W.Woldendorp
  7. Kor. J. Microbiol. v.38 PCR-RFLP and sequence analysis of the rDNA ITS region in the Fusarium spp Lee,Y.M.;Y.K.Choi;B.R.Min
  8. Mol. Biol. Evol. v.9 Rapid surveying of DNA sequence variation in natural populations Lessa,E.P.
  9. Nucleic Acids Res. v.13 Nearly all single base substitutions in DNA fragments joined to a GC-clamp can be detected by denaturing gradient gel electrophoresis Myers,R.M.;S.G.Fischer;L.S.Lerman;T.Maniatis https://doi.org/10.1093/nar/13.9.3131
  10. Nucleic Acids Res. v.13 Modification of the melting properties of duplex DNA by attachment of a GC-rich DNA sequence as determined by denaturing gradient gel electrophoresis Myers,R.M.;S.G.Fischer;T.Maniatis;L.S.Leman https://doi.org/10.1093/nar/13.9.3111
  11. Methods Enzymol v.155 Detection and localization of single base changes by denaturing gradient gel electrophoresis Myers,R.M.;T.Maniatis;L.S.Lerman https://doi.org/10.1016/0076-6879(87)55033-9
  12. FEMS Microbiol. Lett. v.202 ITS region of the rDNA of Pythium longandrum, a new species; its taxonomy and its comparison with related species Paul,B. https://doi.org/10.1111/j.1574-6968.2001.tb10810.x
  13. A revision of the genus Rhizopus no.25 Studies in Mycology Schipper,M.A.A.;J.A.Stalpers;M.A.A.Schipper(ed.);J.A.Stalpers(ed.)
  14. Habilschr. Mucormykosenbei Mensch und Tier. Taxomoie der Erreger. Chemotherapie im Tierexperiment und in der Klinik Scholer,J.J.
  15. J. Gen. Appl. Microbiol. v.46 Genetic similarity among Cercospora apii-group species and their detection in host plant tissue by PCR/RFLP analyses of the rDNA internal transcribed spacer(ITS) Siboe,G.M.;M.Johanne;M.K.Paul https://doi.org/10.2323/jgam.46.69
  16. Amplification and direct sequencing of fungal ribosomal genes form phylogenetics White,T.J.;T.D.Bruns;S.Lee;J.Taylor;M.A.Innis(ed.);D.H.Gelfrand(ed.);J.J.Sninsky(ed.);T.J.White(ed.)