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PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and <t>the</t> <t>wild-type</t> strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.
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PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and <t>the</t> <t>wild-type</t> strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.
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PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and <t>the</t> <t>wild-type</t> strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.
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PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and <t>the</t> <t>wild-type</t> strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.
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PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and <t>the</t> <t>wild-type</t> strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.
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b thuringiensis subsp tenebrionis strain nb 176  (ATCC)
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PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and <t>the</t> <t>wild-type</t> strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.
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Image Search Results


PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and the wild-type strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.

Journal: bioRxiv

Article Title: Development of a CRISPR/Cas9-mediated transformation procedure for the wheat pathogen Zymoseptoria tritici

doi: 10.64898/2026.05.27.728285

Figure Lengend Snippet: PCR amplifications of putative Zymoseptoria tritici CRISPR mutants and the wild-type strain using primers for the target DNA regions and a cartoon of the expected insertion for one mutant as an example. A) M = 10-kb marker ladder (Smartladder); 1 – 3, WT DNA. 1: Mycgr394290 (2,094 bp); 2: Mycgr3107904 (1,621 bp); 3: Mycgr3109710 (1,535 bp); 4: ΔMycgr394290 DNA, amplicon is ∼2,000 bp; 5: ΔMycgr3107904_1 DNA, two DNA fragments of ∼1,500 bp and ∼4,000 bp were obtained; 6: ΔMycgr3107904_2 DNA, amplicon is ∼1,900 bp; 7: ΔMycgr3109710 DNA, amplicon is ∼1,550 bp. B) Schematic representation of the Mycgr3107904 gene disruption by Cas9-mediated transformation coupled with microhomology-directed repair (MDR). The cleavage site of the in vitro -assembled Cas9/sgRNA2, the 60-bp microhomology regions for MDR, the location of the forward and reverse primers for amplification of the target gene, and the expected sizes of each region are represented for the genomic locus of the wild-type IPO323 and the Δ ΔMycgr3107904_1 transformant strain. C) Schematic representation of the expected Mycgr3107904 gene disruption following insertion of the HygB gene.

Article Snippet: Knockout strains of Z. tritici (ΔMycgr3107904_1, ΔMycgr3107904_2, ΔMycgr394290, and ΔMycgr3109710) and the wild-type strain were cultured on PDA supplemented with 100 μg/mL of hygromycin B (GibcoTM, Thermo-Fisher Scientific, Waltham, MA, USA).

Techniques: CRISPR, Mutagenesis, Marker, Amplification, Disruption, Transformation Assay, In Vitro

Disease development of the Δ Mycgr3107904_1 and Δ Mycgr3107904_2 mutants of Zymoseptoria tritici on the susceptible wheat cultivar Taichung 29 compared to wild-type strain IPO323 and mock-inoculated control at A) 14 DPI and B)19 DPI. C) Percentage of symptomatic leaf area of wheat cultivar Taichung29 infected with the Δ Mycgr3107904_1 or Δ Mycgr3107904_2 mutants compared to wild type strain IPO323 at 14 and 19 DPI. Stacked bar plots represent the average of one independent replicated experiment with 3 leaves measured at each time point from each inoculation treatment. Error bars are the standard error of the mean. Bars with different letters at the top indicate significant differences (Tukey’s HSD test, P < 0.05).

Journal: bioRxiv

Article Title: Development of a CRISPR/Cas9-mediated transformation procedure for the wheat pathogen Zymoseptoria tritici

doi: 10.64898/2026.05.27.728285

Figure Lengend Snippet: Disease development of the Δ Mycgr3107904_1 and Δ Mycgr3107904_2 mutants of Zymoseptoria tritici on the susceptible wheat cultivar Taichung 29 compared to wild-type strain IPO323 and mock-inoculated control at A) 14 DPI and B)19 DPI. C) Percentage of symptomatic leaf area of wheat cultivar Taichung29 infected with the Δ Mycgr3107904_1 or Δ Mycgr3107904_2 mutants compared to wild type strain IPO323 at 14 and 19 DPI. Stacked bar plots represent the average of one independent replicated experiment with 3 leaves measured at each time point from each inoculation treatment. Error bars are the standard error of the mean. Bars with different letters at the top indicate significant differences (Tukey’s HSD test, P < 0.05).

Article Snippet: Knockout strains of Z. tritici (ΔMycgr3107904_1, ΔMycgr3107904_2, ΔMycgr394290, and ΔMycgr3109710) and the wild-type strain were cultured on PDA supplemented with 100 μg/mL of hygromycin B (GibcoTM, Thermo-Fisher Scientific, Waltham, MA, USA).

Techniques: Control, Infection