Cloning and characterization of a functional flavanone-3ß-hydroxylase gene from Medicago truncatula.
As a key enzyme in the biosynthesis of flavonols, anthocyanidins and proanthocyanidins, flavanone-3ß-hydroxylase (F3H) plays very important roles in plant stress response. A putative flavanone-3ß-hydroxylase gene from Medicago truncatula (MtF3H), a model legume species, was identified from a bio-data analysis platform. It was speculated to be induced by salt stress based on the outcomes of the analysis platform. The complementary DNA (cDNA) consists of 1499 bp with an open reading frame (ORF) of 1098 bp, which encodes a putative protein of 365 amino acids with a molecular weight of about 41.36 kDa and an isoelectric point of 5.60. To measure the catalytic activity of the protein, the MtF3H gene was ligated to pYES2 vector and heterologously expressed in yeast. The recombinant protein converted naringen into dihydrokaempferol and displayed different enzymatic efficiencies with other flavanones, confirming that MtF3H coding a functional flavanone-3ß-hydroxylase. The expression pattern of the MtF3H gene was analyzed by comparative quantitative RT-PCR and a higher level of expression was observed in the roots than was observed in stems and leaves. Furthermore, the expression was induced by salt stress in the roots, and to a greater extent in the stems, but the response of the gene activity to salt stress in the stems was slower in the first 12 h following treatment when compared to the roots.
Shen
Xiaoye
X
State Key Laboratory for Agro-biotechnology, College of Biological Sciences, China Agricultural University, Beijing, 100193, People's Republic of China. shenxiaoye2008@gmail.com
Martens
Stefan
S
Chen
Mingliang
M
Li
Daofeng
D
Dong
Jiangli
J
Wang
Tao
T
D001711:Biotransformation / Q000187:drug effects / Q000235:genetics; D003001:Cloning, Molecular; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D017343:Genes, Plant / Q000235:genetics*; D046913:Medicago truncatula / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics*; D006899:Mixed Function Oxygenases / Q000235:genetics*; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000187:drug effects / Q000235:genetics; D010802:Phylogeny; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D013312:Stress, Physiological / Q000187:drug effects / Q000235:genetics
D012333:RNA, Messenger; D012965:Sodium Chloride; D006899:Mixed Function Oxygenases; C048797:flavanone 3-dioxygenase