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Journal of Zhejiang University SCIENCE B 2008 Vol.9 No.10 P.811~817

http://doi.org/10.1631/jzus.B0860011


A simple and effective method for total RNA isolation of appressoria in Magnaporthe oryzae


Author(s):  Tong-bao LIU, Jian-ping LU, Xiao-hong LIU, Hang MIN, Fu-cheng LIN

Affiliation(s):  State Key Laboratory for Rice Biology, Biotechnology Institute, Zhejiang University, Hangzhou 310029, China; more

Corresponding email(s):   fuchenglin@zju.edu.cn

Key Words:  Appressorium, Magnaporthe oryzae, RNA isolation


Tong-bao LIU, Jian-ping LU, Xiao-hong LIU, Hang MIN, Fu-cheng LIN. A simple and effective method for total RNA isolation of appressoria in Magnaporthe oryzae[J]. Journal of Zhejiang University Science B, 2008, 9(10): 811~817.

@article{title="A simple and effective method for total RNA isolation of appressoria in Magnaporthe oryzae",
author="Tong-bao LIU, Jian-ping LU, Xiao-hong LIU, Hang MIN, Fu-cheng LIN",
journal="Journal of Zhejiang University Science B",
volume="9",
number="10",
pages="811~817",
year="2008",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B0860011"
}

%0 Journal Article
%T A simple and effective method for total RNA isolation of appressoria in Magnaporthe oryzae
%A Tong-bao LIU
%A Jian-ping LU
%A Xiao-hong LIU
%A Hang MIN
%A Fu-cheng LIN
%J Journal of Zhejiang University SCIENCE B
%V 9
%N 10
%P 811~817
%@ 1673-1581
%D 2008
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B0860011

TY - JOUR
T1 - A simple and effective method for total RNA isolation of appressoria in Magnaporthe oryzae
A1 - Tong-bao LIU
A1 - Jian-ping LU
A1 - Xiao-hong LIU
A1 - Hang MIN
A1 - Fu-cheng LIN
J0 - Journal of Zhejiang University Science B
VL - 9
IS - 10
SP - 811
EP - 817
%@ 1673-1581
Y1 - 2008
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.B0860011


Abstract: 
appressorium formation is an important event in establishing a successful interaction between the rice blast fungus, Magnaporthe oryzae, and its host plant, rice. An understanding of molecular events occurring in appressorium differentiation will give new strategies to control rice blast. A quick and reliable method to extract total RNA from appressorium is essential for studying gene expression during appressorium formation and its mechanism. We found that duplicate film is an efficient substratum for appressorium formation, even when inoculated with high density conidia. When inoculated with conidia at 1×106 ml−1, the percentages of conidium germination and appressorium formation were (97.98±0.67)% and (97.88±0.45)%, respectively. We applied Trizol before appressorium collection for total RNA isolation, and as much as 113.6 μg total RNA was isolated from the mature appressoria at 24 h after inoculation. Functional analysis of two genes, MNH6 and MgATG1, isolated from the cDNA subtractive library, revealed that the quantity of RNA was good enough to construct a cDNA (complementary DNA) library or a cDNA subtractive library. This method may be also applicable for the appressorium RNA isolation of other pathogenic fungi in which conidia differentiate into appressoria in the early stages of host infection.

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article

Reference

[1] Ahn, N., Kim, S., Choi, W., Im, K.H., Lee, Y.H., 2004. Extracellular matrix protein gene, EMP1, is required for appressorium formation and pathogenicity of the rice blast fungus, Magnaporthe grisea. Molecules and Cells, 17(1):166-173.

[2] Balhadère, P.V., Foster, A.J., Talbot, N.J., 1999. Identification of pathogenicity mutants of the rice blast fungus Magnaporthe grisea using insertional mutagenesis. Molecular Plant-Microbe Interactions, 12(2):129-142.

[3] Bourett, T.M., Howard, R.J., 1990. In vitro development of penetration structures in the rice blast fungus Magnaporthe grisea. Canadian Journal of Botany, 68:329-342.

[4] Dean, R.A., 1997. Signal pathways and appressorium morphogenesis. Annual Review of Phytopathology, 35(1):211-234.

[5] Dean, R.A., Talbot, N.J., Ebbole, D.J., Farman, M.L., Mitchell, T.K., Orbach, M.J., Thon, M., Kulkarni, R., Xu, J.R., Pan, H., et al., 2005. The genome sequence of the rice blast fungus Magnaporthe grisea. Nature, 434(7036):980-986.

[6] DeZwaan, T.M., Carroll, A.M., Valent, B., Sweigard, J.A., 1999. Magnaporthe grisea Pth11p is a novel plasma membrane protein that mediates appressorium differentiation in response to inductive substrate cues. Plant Cell, 11(10):2013-2030.

[7] Diatchenko, L., Lau, Y.F., Campbell, A.P., Chenchik, A., Moqadam, F., Huang, B., Lukyanov, S., Lukyanov, K., Gurskaya, N., Sverdlov, E., Siebert, P.D., 1996. Suppression subtractive hybridization: a method for generating differentially regulated or tissue-specific cDNA probes and libraries. Proc. Natl. Acad. Sci. USA, 93(12):6025-6030.

[8] Gilbert, R.D., Johnson, A.M., Dean, R.A., 1996. Chemical signals responsible for appressorium formation in the rice blast fungus Magnaporthe grisea. Physiol. Mol. Plant Pathol., 48(5):335-346.

[9] Gupta, A., Chattoo, B.B., 2007. A novel gene MGA1 is required for appressorium formation in Magnaporthe grisea. Fungal Genet. Biol., 44(11):1157-1169.

[10] Hamer, J.E., Howard, R.J., Chumley, F.G., Valent, B., 1988. A mechanism for surface attachment in spores of a plant pathogenic fungi. Science, 239(4837):288-290.

[11] Hegde, Y., Kolattukudy, P.E., 1997. Cuticular waxes relieve self-inhibition of germination and appressorium formation by the conidia of Magnaporthe grisea. Physiol. Mol. Plant Pathol., 51(2):75-84.

[12] Howard, R.J., Ferrari, M.A., Roach, D.H., Money, N.P., 1991. Penetration of hard substrates by a fungus employing enormous turgor pressures. Proc. Natl. Acad. Sci. USA, 88(24):11281-11284.

[13] Irie, T., Matsumura, H., Terauchi, R., Saitoh, H., 2003. Serial analysis of gene expression (SAGE) of Magnaporthe grisea: genes involved in appressorium formation. Molecular Genetics and Genomics, 270(2):181-189.

[14] Jelitto, T.C., Page, H.A., Read, N.D., 1994. Role of external signals in regulating the pre-penetration phase of infection by the rice blast fungus, Magnaporthe grisea. Planta, 194(4):471-477.

[15] Kamakura, T., Xiao, J.Z., Choi, W.B., Kochi, T., Yamaguchi, S., Teraoka, T., Yamaguchi, I., 1999. cDNA subtractive cloning of genes expressed during early stage of appressoriun formation by Magnaporthe grisea. Biosci. Biotechnol. Biochem., 63(8):1407-1413.

[16] Kamakura, T., Yamaguchi, S., Saitoh, K., Teraoka, T., Yamaguchi, I., 2002. A novel gene, CBP1, encoding a putative extracellular chitin binding protein, may play an important role in the hydrophobic surface sensing of Magnaporthe grisea during appressorium formation. Molecular Plant-Microbe Interactions, 15(5):437-444.

[17] Kim, Y.K., Liu, Z.M., Li, D., Kolattukudy, P.E., 2000. Two novel genes induced by hard-surface contact of Colletotrichum gloeosporioides conidia. J. Bacteriol., 182(17):4688-4695.

[18] Lee, Y.H., Dean, R.A., 1993. cAMP regulates infection structure formation in the plant pathogenic fungus Magnaporthe grisea. Plant Cell, 5(6):693-700.

[19] Lee, Y.H., Dean, R.A., 1994. Hydrophobicity of contact surface induces appressorium formation in Magnaporthe grisea. FEMS Microbiol. Lett., 115(1):71-76.

[20] Liu, X.H., Lu, J.P., Zhang, L., Dong, B., Min, H., Lin, F.C., 2007. Involvement of a Magnaporthe grisea serine/threonine kinase gene, MgATG1, in appressorium turgor and pathogenesis. Eukaryotic Cell, 6(6):997-1005.

[21] Lu, J.P., Liu, T.B., Liu, X.Y., Lin, F.C., 2005a. Representative appressorium stage cDNA library of Magnaporthe grisea. J. Zhejiang Univ. Sci. B, 6(2):132-136.

[22] Lu, J.P., Liu, T.B., Lin, F.C., 2005b. Identification of mature appressorium-enriched transcripts in Magnaporthe grisea, the rice blast fungus, using suppression subtractive hybridization. FEMS Microbiol. Lett., 245(1):131-137.

[23] Lu, J.P., Feng, X.X., Liu, X.H., Lu, Q., Wang, H.K., Lin, F.C., 2007. Mnh6, a nonhistone protein, is required for fungal development and pathogenicity of Magnaporthe grisea. Fungal Genet. Biol., 44(9):819-829.

[24] Ou, S.H., 1985. Rice Diseases, 2nd Ed. Commonwealth Mycological Institute, Kew, UK.

[25] Rauyaree, P., Choi, W., Fang, E., Blackmon, B., Dean, R.A., 2001. Genes expressed during early stages of rice infection with the rice blast fungus Magnaporthe grisea. Molecular Plant Pathology, 2(6):347-354.

[26] Sweigard, J.A., Carroll, A.M., Farrall, L., Chumley, F.G., Valent, B., 1998. Magnaporthe grisea pathogenicity genes obtained through insertional mutagenesis. Molecular Plant-Microbe Interactions, 11(5):404-412.

[27] Talbot, N.J., Ebbole, D.J., Hamer, J.E., 1993. Identification and characterization of MPG1, a gene involved in pathogenicity from the rice blast fungus Magnaporthe grisea. Plant Cell, 5(11):1575-1590.

[28] Xiao, J.Z., Watanabe, T., Kamakura, T., Ohshima, A., Yamaguchi, I., 1994. Studies on cellular differentiation of Magnaporthe grisea: physicochemical aspects of substratum surfaces in relation to appressorium formation. Physiol. Mol. Plant Pathol., 44(3):227-236.

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