title: RolB gene-induced production of isoflavonoids in transformed Maackia amurensis cells.
abstract: Maackia amurensis Rupr. et Maxim is a valuable leguminous tree grown in the Russian Far East, in China, and in Korea. Polyphenols from the heartwood of this species (primarily stilbenes and isoflavonoids) possess strong hepatoprotective activity. Callus culture of M. amurensis produced isoflavonoids and their derivatives. In pharmacological experiments, the callus complex was at least as effective, as the plant complex. To increase the yield of isoflavonoids, calli were transformed with the rolB gene of Agrobacterium rhizogenes. Neomycin phosphotransferase (nptII) gene was used for transgenic cell selection. Three rolB transgenic callus lines with different levels of the rolB gene expression were established. Insertion of the rolB gene caused alterations in callus structure, growth, and isoflavonoid production, and stronger alterations were observed with higher expression levels. MB1, MB2, and MB4 cultures accumulated 1.4, 1.5, and 2.1 % of dry weight (DW) isoflavonoids, respectively. In contrast, the empty vector-transformed MV culture accumulated 1.22 % DW. Isoflavonoid productivity of the obtained MB1, MB2, and MB4 cultures was equal to 117, 112, and 199 mg/L of medium, respectively, comparing to 106 mg/L for the MV culture. High level of expression of the rolB gene in MB4 culture led to a 2-fold increase in the isoflavonoid content and productivity and reliably increased dry biomass accumulation. Lower expression levels of the rolB gene in MB1 and MB2 calli did not significantly enhance biomass accumulation and isoflavonoid content, although the rolB gene activated isoflavonoid biosynthesis during the early growth stages and caused the increased content of several distinct compounds.
authors.affiliation: Institute of Biology and Soil Science, Far East Branch of Russian Academy of Sciences, Stoletija Str., 159, Vladivostok, 690022, Russia. grishchenkoov@gmail.com.
authors.forename: O V
authors.initials: OV
authors.lastname: Grishchenko
authors.affiliation: Institute of Biology and Soil Science, Far East Branch of Russian Academy of Sciences, Stoletija Str., 159, Vladivostok, 690022, Russia.
authors.forename: K V
authors.initials: KV
authors.lastname: Kiselev
authors.affiliation: Institute of Biology and Soil Science, Far East Branch of Russian Academy of Sciences, Stoletija Str., 159, Vladivostok, 690022, Russia.
authors.forename: G K
authors.initials: GK
authors.lastname: Tchernoded
authors.affiliation: Far East Branch of Russian Academy of Sciences, G. V. Elyakov Pacific Institute of Bioorganic Chemistry, Vladivostok, 690022, Russia.
authors.forename: S A
authors.initials: SA
authors.lastname: Fedoreyev
authors.affiliation: Far East Branch of Russian Academy of Sciences, G. V. Elyakov Pacific Institute of Bioorganic Chemistry, Vladivostok, 690022, Russia.
authors.forename: M V
authors.initials: MV
authors.lastname: Veselova
authors.affiliation: Institute of Biology and Soil Science, Far East Branch of Russian Academy of Sciences, Stoletija Str., 159, Vladivostok, 690022, Russia.|Department of Biochemistry, Microbiology and Biotechnology, The School of Natural Sciences, Far Eastern Federal University, Vladivostok, 690950, Russia.
authors.forename: V P
authors.initials: VP
authors.lastname: Bulgakov
authors.affiliation: Institute of Biology and Soil Science, Far East Branch of Russian Academy of Sciences, Stoletija Str., 159, Vladivostok, 690022, Russia.
authors.forename: Y N
authors.initials: YN
authors.lastname: Zhuravlev
mesh_terms: D060054:Agrobacterium / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics*; D018506:Gene Expression Regulation, Plant; D007529:Isoflavones / Q000096:biosynthesis* / Q000737:chemistry; D019868:Kanamycin Kinase / Q000235:genetics; D029901:Maackia / Q000235:genetics* / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000378:metabolism*; D001617:beta-Glucosidase / Q000235:genetics*
chemical_list: D001426:Bacterial Proteins; D007529:Isoflavones; D010940:Plant Proteins; D019868:Kanamycin Kinase; C092241:RolB protein, Agrobacterium rhizogenes; D001617:beta-Glucosidase
keywords: Agrobacterium rhizogenes; Callus culture; Isoflavonoids; Maackia amurensis; Secondary metabolism; rolB