title: Effect of dual Bt-expression transformation vectors on transgene expression in tobacco.
abstract: This study aimed to determine the influence of vector structure on dual Bt gene expression and establish an efficient expression vector using Cry1Ac and Cry3A genes. Four vectors (N4, N5, N10, and S23) were developed and used for genetic transformation of tobacco to obtain insect-resistant transgenic lines. The vectors were constructed using the MAR structure, applying different promoter and enhancer sequences, and changing the transgene open-reading frame sequence. The average Cry1Ac toxalbumin expression quantity was 67 times higher in N5 than in N4 transgenic lines (8.77 and 0.13 μg/g, respectively). In contrast, the average Cry3A toxalbumin expression quantity was 1.5 times higher in N4 than in N5 lines (12.70 and 8.21 μg/g, respectively). The sequences of both Bt genes significantly influenced toxalbumin expression, although upstream Bt genes presented lower expression levels. The average Cry1Ac toxalbumin content was 13 times higher in the transgenic lines of AtADH 5'-non-translated sequence N5 (8.77 mg/g) than in the omega N10 lines (0.67 mg/g). Furthermore, the average Cry1Ac toxalbumin content was 5 times higher in MAR N5 than in non-MAR S23 lines (8.77 and 1.63 mg/g, respectively). The average Cry3A toxalbumin content was 1.3 times higher in N5 than in S23 lines (8.21 and 6.48 mg/g, respectively). Moreover, toxalbumin expression levels differed significantly among the S23-transformed lines. The MAR structure applied on both ends of the genes increased both the level and stability of exogenous gene expression. In conclusion, N5 was the most optimal of the four tested vectors.
authors.affiliation: Institute of Forest Biotechnology, Forestry College, Agricultural University of Hebei, Baoding, China.|Hebei Key Laboratory for Tree Genetic Resources and Forest Protection, Baoding, China.
authors.forename: L N
authors.initials: LN
authors.lastname: Xu
authors.affiliation: Institute of Forest Biotechnology, Forestry College, Agricultural University of Hebei, Baoding, China.|Hebei Key Laboratory for Tree Genetic Resources and Forest Protection, Baoding, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Dong
authors.affiliation: Institute of Forest Biotechnology, Forestry College, Agricultural University of Hebei, Baoding, China.|Hebei Key Laboratory for Tree Genetic Resources and Forest Protection, Baoding, China.
authors.forename: J
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Institute of Forest Biotechnology, Forestry College, Agricultural University of Hebei, Baoding, China.
authors.forename: R X
authors.initials: RX
authors.lastname: Wang
authors.affiliation: Institute of Forest Biotechnology, Forestry College, Agricultural University of Hebei, Baoding, China.
authors.forename: H M
authors.initials: HM
authors.lastname: Liu
authors.affiliation: Institute of Forest Biotechnology, Forestry College, Agricultural University of Hebei, Baoding, China.
authors.forename: Q
authors.initials: Q
authors.lastname: Yang
authors.affiliation: Institute of Forest Biotechnology, Forestry College, Agricultural University of Hebei, Baoding, China yangms100@126.com.|Hebei Key Laboratory for Tree Genetic Resources and Forest Protection, Baoding, China yangms100@126.com.
authors.forename: M S
authors.initials: MS
authors.lastname: Yang
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D000818:Animals; D001413:Bacillus thuringiensis / Q000235:genetics; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D004731:Endotoxins / Q000096:biosynthesis / Q000235:genetics*; D015870:Gene Expression; D005822:Genetic Vectors; D006460:Hemolysin Proteins / Q000096:biosynthesis / Q000235:genetics*; D060434:Herbivory; D007814:Larva / Q000502:physiology; D007915:Lepidoptera / Q000502:physiology*; D010572:Pest Control, Biological; D018515:Plant Leaves / Q000235:genetics* / Q000378:metabolism; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic; D019076:Transgenes
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis