Author:
ZHOU Y Q,ZHANG Y H,WANG W S,WANG X N,DUAN H Y
Abstract
A 1640-bp full-length cDNA of the 3-ketoacyl-CoA thiolase gene was cloned from Rehmannia glutinosa L. leaf using RT-PCR and RACE. Bioinformatics analyses indicated that it contained one 1395-bp open reading frameencoding a thiolase of 464 amino acids with the predicted molecular weight of 49 kDa, and had a similarity of 73-82% to other species at the nucleotide level and 63-88% to other species at the amino acid level, respectively. The recombinant RglKAT was expressed in a bacterial system in an active form, analyzed by PAGE-CBB staining and purified using Ni-agarose chromatography. Real Time quantitative PCR analysis showed that relative expression levels of the RglKATgene differed between R.glutinosa tissues, with the highest levels in the petals and the lowest in the early senescing stems. These results revealed that RglKAT gene could be cloned and heterogeneously expressed, and related to R. glutinosa growth and development.
Publisher
Indian Council of Agricultural Research, Directorate of Knowledge Management in Agriculture
Subject
Agronomy and Crop Science
Reference18 articles.
1. Carrie C, Monika W, Murcha A, Millar H, Smith S M, Whelan J. 2007. Nine 3-ketoacyl-CoA thiolases (KATs) and acetoacetyl- CoA thiolases (ACATs) encoded by five genes in Arabidopsis thaliana are targeted either to peroxisomes or cytosol but not to mitochondria. Plant Molecular Biology 63: 97–108.
2. Dyer J H, Maina A, Gomez I D, Cadet M, Oeljeklaus S and Schiedel A C. 2009. Cloning, expression and purification of an acetoacetyl-CoA thiolase from sunflower cotyledon. International Journal of Bioogical Sciences 5: 736–44.
3. Footitt S, Cornah J E, Pracharoenwattana I, Bryce J H, Smith S M. 2007. The Arabidopsis 3-ketoacyl-CoA thiolase-2, kat2-1, mutant exhibits increased flowering but reduced reproductive success. Journal Experimental Botany 58: 2 959–68.
4. Germain V, Rylott E L, Larson T R, Sherson S M, Bechtold N and Carde J P. 2011. Requirement for 3-ketoacyl-CoA thiolase-2 in peroxisome development, fatty acid -oxidation and breakdown of triacylglycerol in lipid bodies of Arabidopsis seedlings. Plant Journal 28: 1–12.
5. Hou WH, Sun P, Chen QJ and Li XE. 2011. Selection of the reference genes for gene expression studies in Rehmannia glutinosa by Real-time quantitative PCR. Chinese Agricultural Science Bulletin 27: 76–82. Chinese.