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Genome-wide analysis of T-DNA integration into the chromosomes of Magnaporthe oryzae.

Choi J, Park J, Jeon J, Chi MH, Goh J, Yoo SY, Park J, Jung K, Kim H, Park SY, Rho HS, Kim S, Kim BR, Han SS, Kang S, Lee YH - Mol. Microbiol. (2007)

Bottom Line: We identified a total of 1110 T-DNA-tagged locations (TTLs) and processed the resulting data via TAP.Analysis of the TTLs showed that T-DNA integration was biased among chromosomes and preferred the promoter region of genes.Our results support the potential of ATMT as a tool for functional genomics of fungi and show that the TAP is an effective informatics platform for handling data from large-scale insertional mutagenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Agricultural Biotechnology, Center for Fungal Genetic Resources, and Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921, Korea.

ABSTRACT
Agrobacterium tumefaciens-mediated transformation (ATMT) has become a prevalent tool for functional genomics of fungi, but our understanding of T-DNA integration into the fungal genome remains limited relative to that in plants. Using a model plant-pathogenic fungus, Magnaporthe oryzae, here we report the most comprehensive analysis of T-DNA integration events in fungi and the development of an informatics infrastructure, termed a T-DNA analysis platform (TAP). We identified a total of 1110 T-DNA-tagged locations (TTLs) and processed the resulting data via TAP. Analysis of the TTLs showed that T-DNA integration was biased among chromosomes and preferred the promoter region of genes. In addition, irregular patterns of T-DNA integration, such as chromosomal rearrangement and readthrough of plasmid vectors, were also observed, showing that T-DNA integration patterns into the fungal genome are as diverse as those of their plant counterparts. However, overall the observed junction structures between T-DNA borders and flanking genomic DNA sequences revealed that T-DNA integration into the fungal genome was more canonical than those observed in plants. Our results support the potential of ATMT as a tool for functional genomics of fungi and show that the TAP is an effective informatics platform for handling data from large-scale insertional mutagenesis.

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Border truncation, microhomology and host genome deletion at 493 precise junctions. A. Frequency distribution of T-DNA border truncations. B. Distribution of the length of nucleotides identical between genomic and border sequences (microhomology). The expected length of microhomology was plotted on the basis of a random simulation (thick lines). C. Frequency distribution of host genome deletions at 92 insertion sites where sequences at both sides of insertion were determined.
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fig05: Border truncation, microhomology and host genome deletion at 493 precise junctions. A. Frequency distribution of T-DNA border truncations. B. Distribution of the length of nucleotides identical between genomic and border sequences (microhomology). The expected length of microhomology was plotted on the basis of a random simulation (thick lines). C. Frequency distribution of host genome deletions at 92 insertion sites where sequences at both sides of insertion were determined.

Mentions: Left and right T-DNA borders from the selected 493 precise junctions included truncation from the known VirD2 cleavage sites. The LB region was truncated at a high frequency (62%, 101 of 161; Fig. 5A), with the length of deletions ranging from 1 to 147 bp. On the other hand, the RB region appeared well preserved (4% 13 of 332; Fig. 5A), with the length of deletions ranging from 1 to 74 bp. Small truncations with less than 35 bp represented 97.0% and 84.6% of the LB and RB truncations respectively, with only a few truncations exceeding 35 bp at both ends of the T-DNA (Fig. 5A).


Genome-wide analysis of T-DNA integration into the chromosomes of Magnaporthe oryzae.

Choi J, Park J, Jeon J, Chi MH, Goh J, Yoo SY, Park J, Jung K, Kim H, Park SY, Rho HS, Kim S, Kim BR, Han SS, Kang S, Lee YH - Mol. Microbiol. (2007)

Border truncation, microhomology and host genome deletion at 493 precise junctions. A. Frequency distribution of T-DNA border truncations. B. Distribution of the length of nucleotides identical between genomic and border sequences (microhomology). The expected length of microhomology was plotted on the basis of a random simulation (thick lines). C. Frequency distribution of host genome deletions at 92 insertion sites where sequences at both sides of insertion were determined.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169514&req=5

fig05: Border truncation, microhomology and host genome deletion at 493 precise junctions. A. Frequency distribution of T-DNA border truncations. B. Distribution of the length of nucleotides identical between genomic and border sequences (microhomology). The expected length of microhomology was plotted on the basis of a random simulation (thick lines). C. Frequency distribution of host genome deletions at 92 insertion sites where sequences at both sides of insertion were determined.
Mentions: Left and right T-DNA borders from the selected 493 precise junctions included truncation from the known VirD2 cleavage sites. The LB region was truncated at a high frequency (62%, 101 of 161; Fig. 5A), with the length of deletions ranging from 1 to 147 bp. On the other hand, the RB region appeared well preserved (4% 13 of 332; Fig. 5A), with the length of deletions ranging from 1 to 74 bp. Small truncations with less than 35 bp represented 97.0% and 84.6% of the LB and RB truncations respectively, with only a few truncations exceeding 35 bp at both ends of the T-DNA (Fig. 5A).

Bottom Line: We identified a total of 1110 T-DNA-tagged locations (TTLs) and processed the resulting data via TAP.Analysis of the TTLs showed that T-DNA integration was biased among chromosomes and preferred the promoter region of genes.Our results support the potential of ATMT as a tool for functional genomics of fungi and show that the TAP is an effective informatics platform for handling data from large-scale insertional mutagenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Agricultural Biotechnology, Center for Fungal Genetic Resources, and Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921, Korea.

ABSTRACT
Agrobacterium tumefaciens-mediated transformation (ATMT) has become a prevalent tool for functional genomics of fungi, but our understanding of T-DNA integration into the fungal genome remains limited relative to that in plants. Using a model plant-pathogenic fungus, Magnaporthe oryzae, here we report the most comprehensive analysis of T-DNA integration events in fungi and the development of an informatics infrastructure, termed a T-DNA analysis platform (TAP). We identified a total of 1110 T-DNA-tagged locations (TTLs) and processed the resulting data via TAP. Analysis of the TTLs showed that T-DNA integration was biased among chromosomes and preferred the promoter region of genes. In addition, irregular patterns of T-DNA integration, such as chromosomal rearrangement and readthrough of plasmid vectors, were also observed, showing that T-DNA integration patterns into the fungal genome are as diverse as those of their plant counterparts. However, overall the observed junction structures between T-DNA borders and flanking genomic DNA sequences revealed that T-DNA integration into the fungal genome was more canonical than those observed in plants. Our results support the potential of ATMT as a tool for functional genomics of fungi and show that the TAP is an effective informatics platform for handling data from large-scale insertional mutagenesis.

Show MeSH
Related in: MedlinePlus