Transformation of carotenoid biosynthetic genes using a micro-cross section method in kiwifruit (Actinidia deliciosa cv. Hayward).
Genetic transformation using a micro-cross section (MCS) technique was conducted to improve the carotenoid content in kiwifruit (Actinidia deliciosa cv. Hayward). The introduced carotenoid biosynthetic genes include geranylgeranyl diphosphate synthase (GGPS), phytoene desaturase (PDS), ζ-carotene desaturase (ZDS), β-carotene hydroxylase (CHX), and phytoene synthase (PSY). The transformed explants were selected on half-strength MS medium containing 0.001 mg l(-1) of 2,4-D and 0.1 mg l(-1) of zeatin, either 5 mg l(-1) hygromycin or 25 mg l(-1) kanamycin, and 500 mg l(-1) cefotaxime. The genomic PCR, genomic Southern blot analysis, and RT-PCR were performed to confirm the integration and expression of the transgenes. The transformation efficiencies of either kanamycin- or hygromycin-resistant shoots ranged from 2.9 to 22.1% depending on the target genes, and from 2.9 to 24.2% depending on the reporter genes. The selection efficiencies ranged from 66.7 to 100% for the target genes and from 95.8 to 100% for the reporter genes. Changes of carotenoid content in the several PCR-positive plants were determined by UPLC analysis. As a result, transgenic plants expressing either GGPS or PSY increased about 1.2- to 1.3-fold in lutein or β-carotene content compared to non-transgenic plants. Our results suggest that the Agrobacterium-mediated transformation efficiency of kiwifruit can be greatly increased by this MCS method and that the carotenoid biosynthetic pathway can be modified in kiwifruit by genetic transformation. Our results further suggest that GGPS and PSY genes could be major target genes to increase carotenoid contents in kiwifruit.
Kim
Misun
M
Agricultural Research Center for Climate Change, National Institute of Horticultural and Herbal Science, Rural Development Administration, Jeju, Korea.
Kim
Seong-Cheol
SC
Song
Kwan Jeong
KJ
Kim
Ho Bang
HB
Kim
In-Jung
IJ
Song
Eun-Young
EY
Chun
Seung-Jong
SJ
D029042:Actinidia / Q000235:genetics*; D000900:Anti-Bacterial Agents / Q000008:administration & dosage; D001483:Base Sequence; D002338:Carotenoids / Q000096:biosynthesis*; D002853:Chromatography, Liquid; D017931:DNA Primers; D017343:Genes, Plant; D030821:Plants, Genetically Modified / Q000235:genetics; D016133:Polymerase Chain Reaction; D014170:Transformation, Genetic
D000900:Anti-Bacterial Agents; D017931:DNA Primers; D002338:Carotenoids