Ssu72 Regulates Fungal Development, Aflatoxin Biosynthesis and Pathogenicity in Aspergillus flavus
Abstract
:1. Introduction
2. Results
2.1. Identification of Putative Phosphatase Ssu72 in A. flavus
2.2. Construction of the ssu72 Mutant Strains
2.3. Ssu72 Is Involved in Vegetative Growth in A. flavus
2.4. Ssu72 Is Critical for Conidiation in A. flavus
2.5. Ssu72 Is Required for Sclerotia Formation in A. flavus
2.6. Ssu72 Plays a Positive Role in Regulation of Aflatoxin Biosynthesis
2.7. Ssu72 Contributes to Pathogenicity in Crop Seeds in A. flavus
2.8. Ssu72 Response to Multiple Stresses in A. flavus
3. Discussion
4. Materials and Methods
4.1. Strains and Culture Conditions
4.2. Mutant Strains Construction
4.3. Phenotpic Assays
4.4. Aflatoxin Production Assays
4.5. Pathogenicity Assays
4.6. Stress Assay
4.7. RNA Extraction and Quantitative Real-Time PCR Analysis
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Strains | Genotype Description | Reference |
---|---|---|
A. flavus CA14 PTS | ∆ku70, ∆pyrG | Our lab |
wild-type (WT) | ∆ku70, ∆pyrG::AfpyrG | This study |
∆ssu72 | ∆ku70, ∆pyrG::AfpyrG, ∆ssu72 | This study |
∆ssu72-Com | ∆ku70, ∆ssu72::Aflssu72, ∆pyrG::AfpyrG | This study |
Primer Name | Sequence (5′–3′) | Application |
---|---|---|
ssu72-AF | AAACCGACCACGAAGACA | 5 ‘UTR of ssu72 |
ssu72-AR | GGGTGAAGAGCATTGTTTGAGGCTCAAGGAGGCTGGAAGAT | |
ssu72-BF | GCATCAGTGCCTCCTCTCAGACGGCTAGGGTCAACGAACA | 3′UTR of ssu72 |
ssu72-BR | CCCTTCCCTCCTTCAGCA | |
pyrG-F | GCCTCAAACAATGCTCTTCACCC | fumigatus pyrG |
pyrG-R | GTCTGAGAGGAGGCACTGATGC | |
ssu72-NF | GGTCCACTGGGTGGTAAT | Fusion PCR |
ssu72-NR | GCACGATACAAGGCGATGG | |
ssu72-OF | ACCCAGGAGCAACAGTCA | ssu72 ORF verification |
ssu72-OR | CCAGCCTTCAGAGTTATTCG | |
P801-R | CAGGAGTTCTCGGGTTGTCG | Verification of AP and BP |
P1020-F | CAGAGTATGCGGCAAGTCA | |
ssu72-Com-AF | AACTAGTGAACACATCTTC | 5′UTR of ∆ssu72-Com |
ssu72-Com-AR | GGGTGAAGAGCATTGTTTGAGGCCCAATGTTCGTTGACCCTA | |
ssu72-Com-BF | GCATCAGTGCCTCCTCTCAGACCTGTATGCAGATCCAAAT | 3′UTR of ∆ssu72-Com |
ssu72-Com-BR | CTTCTAAAGCTCCTATCC | |
ssu72-Com-NF | TTCTGTTGGCCTGCGTAT | Fusion PCR |
ssu72-Com-NR | GTATGCCTCTTGACTCCC |
Primer Name | Sequence (5′–3′) | Application |
---|---|---|
ssu72-F | GAGTCTTCAGACGGGACTGC | ssu72 detection |
ssu72-R | CACATTAGGTTGCGTGATGG | |
brlA-F | GCCTCCAGCGTCAACCTTC | brlA qRT-PCR |
brlA-R | TCTCTTCAAATGCTCTTGCCTC | |
abaA-F | TCTTCGGTTGATGGATGATTTC | abaA qRT-PCR |
abaA-R | CCGTTGGGAGGCTGGGT | |
nsdC-F | GCCAGACTTGCCAATCAC | nsdC qRT-PCR |
nsdC-R | CATCCACCTTGCCCTTTA | |
nsdD-F | GGACTTGCGGGTCGTGCTA | nsdD qRT-PCR |
nsdD-R | AGAACGCTGGGTCTGGTGC | |
aflR-F | AAAGCACCCTGTCTTCCCTAAC | aflR qRT-PCR |
aflR-R | GAAGAGGTGGGTCAGTGTTTGTAG | |
aflS-F | CGAGTCGCTCAGGCGCTCAA | aflS qRT-PCR |
aflS-R | GCTCAGACTGACCGCCGCTC | |
aflK-F | GAGCGACAGGAGTAACCGTAAG | aflK qRT-PCR |
aflK-R | CCGATTCCAGACACCATTAGCA | |
aflQ-F | GTCGCATATGCCCCGGTCGG | aflQ qRT-PCR |
aflQ-R | GGCAACCAGTCGGGTTCCGG | |
aflO-F | GATTGGGATGTGGTCATGCGATT | aflO qRT-PCR |
aflO-R | GCCTGGGTCCGAAGAATGC | |
aflP-F | ACGAAGCCACTGGTAGAGGAGATG | aflP qRT-PCR |
aflP-R | GTGAATGACGGCAGGCAGGT | |
actin-F | ACGGTGTCGTCACAAACTGG | The endogenos gene |
actin-R | CGGTTGGACTTAGGGTTGATAG |
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Yang, G.; Cao, X.; Qin, L.; Yan, L.; Hong, R.; Yuan, J.; Wang, S. Ssu72 Regulates Fungal Development, Aflatoxin Biosynthesis and Pathogenicity in Aspergillus flavus. Toxins 2020, 12, 717. https://doi.org/10.3390/toxins12110717
Yang G, Cao X, Qin L, Yan L, Hong R, Yuan J, Wang S. Ssu72 Regulates Fungal Development, Aflatoxin Biosynthesis and Pathogenicity in Aspergillus flavus. Toxins. 2020; 12(11):717. https://doi.org/10.3390/toxins12110717
Chicago/Turabian StyleYang, Guang, Xiaohong Cao, Ling Qin, Lijuan Yan, Rongsheng Hong, Jun Yuan, and Shihua Wang. 2020. "Ssu72 Regulates Fungal Development, Aflatoxin Biosynthesis and Pathogenicity in Aspergillus flavus" Toxins 12, no. 11: 717. https://doi.org/10.3390/toxins12110717
APA StyleYang, G., Cao, X., Qin, L., Yan, L., Hong, R., Yuan, J., & Wang, S. (2020). Ssu72 Regulates Fungal Development, Aflatoxin Biosynthesis and Pathogenicity in Aspergillus flavus. Toxins, 12(11), 717. https://doi.org/10.3390/toxins12110717