PMC6690219_01
The accuracy of TDNAscan and similar tools with different coverage simulated data. The green, orange, blue, and yellow bars represent the F-scores of TDNAscan, ITIS, BreakDancer, and DELLY, respectively.
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Compared with these 18 T-DNA insertions, we successfully identified 17 T-DNA and additional truncated T-DNA insertions ( Figure 3A  and  Supplementary File 2 ).
The graph of case studies. (A). Venn diagram of identified T-DNA insertions in published data. (B). PCR validation of predicted T-DNA insertion at location 2465576, the causative insertion of hlb3. (C). PCR confirmation of the remaining two insertion sites to show further validation of the TDNAscan prediction.
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Compared with these 18 T-DNA insertions, we successfully identified 17 T-DNA and additional truncated T-DNA insertions ( Figure 3A  and  Supplementary File 2 ). We were able to amplify the three chimeric products and verify the junctions between the T-DNA and genomic DNA via sequencing ( Figure 3B, C ).
The Arabidopsis HLB3 gene encodes the class II formin AtFH20. (A-B) The hlb3-1/atfh20-1 and hlb3-2/atfh20-2 mutants exhibit heightened sensitivity to the root growth inhibitory effects of latrunculin B when compared to wild type (arrows in B). Four-day-old seedlings of all genotypes were transferred to plates containing 50 nM latrunculin B and imaged after 3 days. (C) Quantification of primary root displacement after transfer to latrunculin B. Statistical significance was determined by one-way ANOVA. Means (n > 40) +- SE. Different letters indicate significant differences among means (P < 0.05, Tukey's test). (D) Schematic diagram of the HLB3/AtFH20 (AT5G07740) gene. Black boxes indicate exons, and lines indicate introns. Inverted triangles show the position of T-DNA for hlb3-1/atfh20-1 and hlb3-2/atfh20-2.
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TDNAscan can be used to identify the disrupted gene in the Arabidopsis hypersensitive to latrunculin B (hlb3) mutant. The recessive hlb3 mutant exhibits heightened sensitivity to the actin-disrupting drug, latrunculin B, and was isolated from the same T-DNA mutagenized population as in our previously published hlb1mutant ( Figure 4A-C ). AT5G07740 ( Figure 4D ) and intron 7 of AT5G01100. AT5G07740 is annotated to encode a FORMIN Homology 20 (AtFH20) protein, while AT5G01100 encodes a O-fucosyltransferase family protein. We verified that SALK_137002 had a T-DNA insertion in the sixth exon of AT5G07740 ( Figure 4D ). B compared to wild type, indicating that the HLB3 is AtFH20 ( Figure 4A-C ). Based on our findings, the hlb3 was renamed hlb3-1/Atfh20-1and SALK_137002 as hlb3-2/Atfh20-2 ( Figure 4D ).