Full Text:   <3092>

Summary:  <1702>

Suppl. Mater.: 

CLC number: S436.661.1+1

On-line Access: 2014-01-28

Received: 2013-08-12

Revision Accepted: 2013-12-06

Crosschecked: 2014-01-17

Cited: 3

Clicked: 6054

Citations:  Bibtex RefMan EndNote GB/T7714

-   Go to

Article info.
Open peer comments

Journal of Zhejiang University SCIENCE B 2014 Vol.15 No.2 P.116-124


Improvement of a gene targeting system for genetic manipulation in Penicillium digitatum * #

Author(s):  Qian Xu, Cong-yi Zhu, Ming-shang Wang, Xue-peng Sun, Hong-ye Li

Affiliation(s):  . Institute of Biotechnology, Zhejiang University, Hangzhou 310058, China

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

Key Words:  Penicillium digitatum , Efficiency, Gene targeting, Non-homologous end-joining (NHEJ) pathway, Ku80

Qian Xu, Cong-yi Zhu, Ming-shang Wang, Xue-peng Sun, Hong-ye Li. Improvement of a gene targeting system for genetic manipulation in Penicillium digitatum[J]. Journal of Zhejiang University Science B, 2014, 15(2): 116-124.

@article{title="Improvement of a gene targeting system for genetic manipulation in Penicillium digitatum",
author="Qian Xu, Cong-yi Zhu, Ming-shang Wang, Xue-peng Sun, Hong-ye Li",
journal="Journal of Zhejiang University Science B",
publisher="Zhejiang University Press & Springer",

%0 Journal Article
%T Improvement of a gene targeting system for genetic manipulation in Penicillium digitatum
%A Qian Xu
%A Cong-yi Zhu
%A Ming-shang Wang
%A Xue-peng Sun
%A Hong-ye Li
%J Journal of Zhejiang University SCIENCE B
%V 15
%N 2
%P 116-124
%@ 1673-1581
%D 2014
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B1300213

T1 - Improvement of a gene targeting system for genetic manipulation in Penicillium digitatum
A1 - Qian Xu
A1 - Cong-yi Zhu
A1 - Ming-shang Wang
A1 - Xue-peng Sun
A1 - Hong-ye Li
J0 - Journal of Zhejiang University Science B
VL - 15
IS - 2
SP - 116
EP - 124
%@ 1673-1581
Y1 - 2014
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.B1300213

Penicillium digitatum is the most important pathogen of postharvest citrus. gene targeting can be done in P. digitatum using homologous recombination via Agrobacterium tumefaciens mediated transformation (ATMT), but the frequencies are often very low. In the present study, we replaced the Ku80 homolog (a gene of the non-homologous end-joining (NHEJ) pathway) with the hygromycin resistance cassette (hph) by ATMT. No significant change in vegetative growth, conidiation, or pathogenicity was observed in Ku80-deficient strain (ΔPdKu80) of P. digitatum. However, using ΔPdKu80 as a targeting strain, the gene-targeting frequencies for both genes PdbrlA and PdmpkA were significantly increased. These results suggest that Ku80 plays an important role in homologous integration and the created ΔPdKu80 strain would be a good candidate for rapid gene function analysis in P. digitatum.




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


[1] Bateman, A., Coin, L., Durbin, R., 2004. The Pfam protein families database. Nucl Acids Res, 32(Suppl. 1):D138-D141. 

[2] Chang, P.K., 2008. A highly efficient gene-targeting system for Aspergillus parasiticusLett Appl Microbiol, 46(5):587-592. 

[3] Choquer, M., Robin, G., Le Pêcheur, P., 2008.  Ku70 or Ku80 deficiencies in the fungus Botrytis cinerea facilitate targeting of genes that are hard to knock out in a wild-type context. FEMS Microbiol Lett, 289(2):225-232. 

[4] da Silva Ferreira, M.E., Kress, M.R., Savoldi, M., 2006. The akuB KU80 mutant deficient for nonhomologous end joining is a powerful tool for analyzing pathogenicity in Aspergillus fumigatusEukaryot Cell, 5(1):207-211. 

[5] Eckert, J.W., Eaks, I., 1989. Postharvest disorders and diseases of citrus fruits. Citrus Ind, 5:179-260. 

[6] Haber, J.E., 2000. Partners and pathways: repairing a double-strand break. Trends Genet, 16(6):259-264. 

[7] Hamamoto, H., Hasegawa, K., Nakaune, R., 2000. Tandem repeat of a transcriptional enhancer upstream of the sterol 14α-demethylase gene (CYP51) in Penicillium digitatumAppl Environ Microbiol, 66(8):3421-3426. 

[8] Ishibashi, K., Suzuki, K., Ando, Y., 2006. Nonhomologous chromosomal integration of foreign DNA is completely dependent on MUS-53 (human Lig4 homolog) in NeurosporaPNAS, 103(40):14871-14876. 

[9] Jiang, J., Liu, X., Yin, Y., 2011. Involvement of a velvet protein FgVeA in the regulation of asexual development, lipid and secondary metabolisms and virulence in Fusarium graminearumPLoS ONE, 6(11):e28291

[10] Jones, J.M., Gellert, M., Yang, W., 2001. A Ku bridge over broken DNA. Structure, 9(10):881-884. 

[11] Kanaar, R., Hoeijmakers, J.H., van Gent, D.C., 1998. Molecular mechanisms of DNA double-strand break repair. Trends Cell Biol, 8(12):483-489. 

[12] Kanetis, L., Frster, H., Adaskaveg, J.E., 2007. Comparative efficacy of the new postharvest fungicides azoxystrobin, fludioxonil, and pyrimethanil for managing citrus green mold. Plant Dis, 91(11):1502-1511. 

[13] Krappmann, S., Sasse, C., Braus, G.H., 2006. Gene targeting in Aspergillus fumigatus by homologous recombination is facilitated in a nonhomologous end-joining-deficient genetic background. Eukaryot Cell, 5(1):212-215. 

[14] Li, Z.H., Du, C.M., Zhong, Y.H., 2010. Development of a highly efficient gene targeting system allowing rapid genetic manipulations in Penicillium decumbensAppl Microbiol Biotechnol, 87(3):1065-1076. 

[15] Macarisin, D., Cohen, L., Eick, A., 2007.  Penicillium digitatum suppresses production of hydrogen peroxide in host tissue during infection of citrus fruit. Phytopathology, 97(11):1491-1500. 

[16] Marcet-Houben, M., Ballester, A.R., de la Fuente, B., 2012. Genome sequence of the necrotrophic fungus Penicillium digitatum, the main postharvest pathogen of citrus. BMC Genomics, 13(1):646

[17] Maruyama, J.I., Kitamoto, K., 2008. Multiple gene disruptions by marker recycling with highly efficient gene-targeting background (∆ligD) in Aspergillus oryzaeBiotechnol Lett, 30(10):1811-1817. 

[18] Meyer, V., Arentshorst, M., El-Ghezal, A., 2007. Highly efficient gene targeting in the Aspergillus niger kusA mutant. J Biotechnol, 128(4):770-775. 

[19] Nakaune, R., Hamamoto, H., Imada, J., 2002. A novel ABC transporter gene, PMR5, is involved in multidrug resistance in the phytopathogenic fungus Penicillium digitatumMol Genet Genomics, 267(2):179-185. 

[20] Nayak, T., Szewczyk, E., Oakley, C.E., 2006. A versatile and efficient gene-targeting system for Aspergillus nidulansGenetics, 172(3):1557-1566. 

[21] Ninomiya, Y., Suzuki, K., Ishii, C., 2004. Highly efficient gene replacements in Neurospora strains deficient for nonhomologous end-joining. PNAS, 101(33):12248-12253. 

[22] Sun, X., Ruan, R., Lin, L., 2013. Genomewide investigation into DNA elements and ABC transporters involved in imazalil resistance in Penicillium digitatumFEMS Microbiol Lett, 348(1):11-18. 

[23] van Dyck, E., Stasiak, A.Z., Stasiak, A., 1999. Binding of double-strand breaks in DNA by human Rad52 protein. Nature, 398(6729):728-731. 

[24] Villalba, F., Collemare, J., Landraud, P., 2008. Improved gene targeting in Magnaporthe grisea by inactivation of MgKU80 required for non-homologous end joining. Fungal Genet Biol, 45(1):68-75. 

[25] Wang, J.Y., Li, H.Y., 2008. Agrobacterium tumefaciens-mediated genetic transformation of the phytopathogenic fungus Penicillium digitatumJ Zhejiang Univ-Sci B, 9(10):823-828. 

[26] Wang, J.Y., Sun, X.P., Lin, L.Y., 2012. PdMfs1, a major facilitator superfamily transporter from Penicillium digitatum, is partially involved in the imazalil-resistance and pathogenicity. Afr J Microbiol Res, 6(1):95-105. 

[27] Wang, N.Y., Yang, S.L., Lin, C.H., 2011. Gene inactivation in the citrus pathogenic fungus Alternaria alternata defect at the Ku70 locus associated with non-homologous end joining. World J Microbiol Biotechnol, 27(8):1817-1826. 

[28] Zhang, T., Xu, Q., Sun, X., 2013. The calcineurin-responsive transcription factor Crz1 is required for conidation, full virulence and DMI resistance in Penicillium digitatumMicrobiol Res, 168(4):211-222. 

[29] Zhang, T., Sun, X., Xu, Q., 2013.  PdSNF1, a sucrose non-fermenting protein kinase gene, is required for Penicillium digitatum conidiation and virulence. Appl Microbiol Biotechnol, 97(12):5433-5445. 

[30] Zhang, T., Sun, X., Xu, Q., 2013. The pH signaling transcription factor PacC is required for full virulence in Penicillium digitatumAppl Microbiol Biotechnol, 97(20):9087-9098. 

[31] Zhang, Z., Zhu, Z., Ma, Z., 2009. A molecular mechanism of azoxystrobin resistance in Penicillium digitatum UV mutants and a PCR-based assay for detection of azoxystrobin-resistant strains in packing- or store-house isolates. Int J Food Microbiol, 131(2-3):157-161. 

Open peer comments: Debate/Discuss/Question/Opinion


Qing-qing Huang<819075736@qq.com>

2015-06-12 10:12:23

Dear author,
Where is the Table S1,I couldn't find it. I want to make it as a reference.I am looking forward to your help

Please provide your name, email address and a comment

Journal of Zhejiang University-SCIENCE, 38 Zheda Road, Hangzhou 310027, China
Tel: +86-571-87952783; E-mail: cjzhang@zju.edu.cn
Copyright © 2000 - 2023 Journal of Zhejiang University-SCIENCE