High-efficiency Agrobacterium rhizogenes-mediated transformation of heat inducible sHSP18.2-GUS in Nicotiana tabacum.
The chimerical gene, Arabidopsis thaliana sHSP18.2 promoter fused to E. coli gusA gene, was Agrobacterium rhizogenes-mediated transformed into Nicotiana tabacum as a heat-regulatable model, and the thermo-inducible expression of GUS activity in N. tabacum transgenic hairy roots was profiled. An activation of A. rhizogenes with acetosyringone (AS) before cocultured with tobacco's leaf disc strongly promoted transgenic hairy roots formation. Transgenic hairy roots formation efficiency of A. rhizogenes precultured with 200 microM AS supplementation was 3.1-fold and 7.5-fold, respectively, compared to the formation efficiency obtained with and without AS supplementation in coculture. Transgenic hairy roots transformed with different AS concentration exhibited a similar pattern of thermo-inducibility after 10 min to 3 h heat treatments detected by GUS expression. The peak of expressed GUS specific activity, 399,530 pmol MUG per mg total protein per min, of the transgenic hairy roots was observed at 48 h after 3 h of 42 degrees C heat treatment, and the expressed GUS specific activity was 7-26 times more than that reported in A. thaliana, tobacco BY-2 cells and Nicotiana plumbaginifolia. Interference caused by AS supplementation on the growth of transgenic hairy roots, time-course of GUS expression and its expression level were not observed.
Chen
Shih-Cheng
SC
Graduate Institute of Microbiology and Biochemistry, National Taiwan University, 1, Sec. 4, Roosevelt Road, Taipei, Taiwan.
Liu
Hui-Wen
HW
Lee
Kung-Ta
KT
Yamakawa
Takashi
T
D000098:Acetophenones / Q000494:pharmacology; D029681:Arabidopsis Proteins / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism*; D050888:Heat-Shock Proteins, Small / Q000235:genetics*; D006358:Hot Temperature; D018517:Plant Roots / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012231:Rhizobium / Q000235:genetics*; D013997:Time Factors; D014026:Nicotiana / Q000235:genetics*; D014170:Transformation, Genetic
D000098:Acetophenones; D029681:Arabidopsis Proteins; D029968:Escherichia coli Proteins; D050888:Heat-Shock Proteins, Small; D011993:Recombinant Fusion Proteins; C051667:acetosyringone; D005966:Glucuronidase