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KMID : 0545120130230010022
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Journal of Microbiology and Biotechnology
2013 Volume.23 No. 1 p.22 ~ p.28
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Homologous Expression and T3SS-Dependent Secretion of TAP-Tagged Xo2276 in Xanthomonas oryzae pv. oryzae Induced by Rice Leaf Extract and Its Direct In Vitro Recognition of Putative Target DNA Sequence
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Kim Seung-Hwan
Thi-Dieu-Hanh Nguyen
Lee Joo-Hee
Hong Myoung-Ki
Tan-Viet Pham
Ahn Yeh-Jin
Lee Byoung-Moo
Han Ye-Sun
Kim Dong-Eun
Kim Jeong-Gu
Kang Lin-Woo
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Abstract
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Xo2276 is a putative transcription activator-like effector (TALE) in Xanthomonas oryzae pv. oryzae (Xoo). Xo2276 was expressed with a TAP-tag at the C-terminus in Xoo cells to enable quantitative analysis of protein expression and secretion. Nearly all TAP-tagged Xo2276 existed in an insoluble form; addition of rice leaf extracts from a Xoosusceptible rice cultivar, Milyang23, significantly stimulated secretion of TAP-tagged Xo2276 into the medium. In a T3SS-defective Xoo mutant strain, secretion of TAPtagged Xo2276 was blocked. Xo2276 is a Xoo ortholog of Xanthomonas campestris pv. vesicatoria (Xcv) AvrBs3 and contains a conserved DNA-binding domain (DBD), which includes 19.5 tandem repeats of 34 amino acids. Xo2276- DBD was expressed in E. coli and purified. Direct in vitro recognition of Xo2276-DBD on a putative target DNA sequence was confirmed using an electrophoretic mobility shift assay. This is the first study measuring the homologous expression and secretion of Xo2276 in vitro using rice leaf extract and its direct in vitro binding to the specific target DNA sequence.
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KEYWORD
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AvrBs3, TALE, DNA-binding domain, Xanthomonas oryzae pv. oryzae
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