Journal: Eukaryotic Cell

Article Title: Use of the Plant Defense Protein Osmotin To Identify Fusarium oxysporum Genes That Control Cell Wall Properties ▿ Genes That Control Cell Wall Properties ▿ ‡

doi: 10.1128/EC.00316-09

Figure Lengend Snippet: Disruption of the FOR3 gene in F. oxysporum f. sp. nicotianae . (A) Schematic view of the FOR3 gene disruption strategy. The DNA inserts of p FOR3 and p for3 ::HYG are shown. The FOR3 coding region is indicated by the gray striped box and noncoding regions by the gray line. The vector sequences are indicated by the black line. Restriction sites on the cloning vector are indicated in black, and those in FOR3 are indicated in gray. The sizes of cloned DNA fragments (double-headed arrows), as well as the probes used for Southern (dotted line) or Northern (dashed line) blot analyses, are indicated. Restriction site abbreviations are as follows: E, EcoRV; E1, EcoRI; H, HindIII; K, KpnI; P, PstI; S, SalI; Xb, XbaI; Xh, XhoI. (B) Southern blot of restriction-digested genomic DNA of the wild-type ( FOR3 ) and Δ for3 mutant strains. Restriction site abbreviations are as in panel A. (C) Northern blot of total RNA of the wild-type ( FOR3 ) and Δ for3 mutant strains with a FOR3 probe ( FOR3 ) and the corresponding ethidium bromide signal (RNA) shown.

Article Snippet: To construct p for3 ::HYG, a 0.7-kb fragment containing the 3′ part of the FOR3 ORF (+3205 bp to +3941 bp) was amplified from F. oxysporum genomic DNA by PCR using the primer pair 5′-ATCGAGTCTTGCCGACGAT-3′/5′-TGAGATCCGTCTTCAGGATC-3′ and inserted into the EcoRV site of the pSTblue-1 vector (Novagen, Madison, WI).

Techniques: Plasmid Preparation, Clone Assay, Northern Blot, Southern Blot, Mutagenesis