title: Effects of BmKIT 3 R gene transfer on pupal development of Bombyx mori Linnaeus using a Gal4/UAS binary transgenic system.
abstract: The pupal stage of the silkworm  Bombyx mori  Linnaeus lasts for approximately two weeks. However, prolongation of pupal duration would reduce the labor required to process and dry fresh cocoons. This study investigated the effects of BmKIT(3)(R) gene (from the Chinese scorpion  Buthus martensii  Karsch) transfer on the pupal development of  B. mori  using a Gal4/UAS binary transgenic system. Gal4 driven by a pupa-specific promoter BmWCP4 (from a  B. mori  wing-cuticle protein gene) or PDP (from a  B. mori  cocoonase gene), and BmKIT(3)(R) driven by a UAS cis-acting element were used to construct novel piggyBac-derived plasmids containing a neomycin-resistance gene (neo) controlled by the  Bombyx mori  nucleopolyhedrovirus (BmNPV) ie-1 (immediate-early gene) promoter and a green fluorescent protein gene (gfp) under the control of the  B. mori  actin 3 (A3) promoter. The vector was transferred into silkworm eggs by sperm-mediated gene transfer. Transgenic silkworms were produced after screening for neo and gfp genes, and gene transfer was verified by polymerase chain reaction and dot-blot hybridization. The larval development of the hybrid progeny of Gal4- and UAS-transgenic silkworms was similar to that of normal silkworms, but some pupae failed to metamorphose into moths, and the development of surviving pupae was arrested as a result of BmKIT(3)(R) expression. Moreover, Gal4 driven by the BmWCP4 promoter delayed pupal development more effectively than that driven by the PDP promoter in the Gal4/UAS binary transgenic system. Pupal durations of hybrid transgenic silkworm progeny with BmWCP4 and PDP promoters were approximately 5, 2, and 4 days longer, respectively, compared to corresponding normal silkworms, BmWCP4/Gal4, and UAS/BmKIT(3)(R) transgenic silkworms, respectively. These results suggest new avenues of research for prolonging the pupal duration of silkworms.
authors.affiliation: School of Biology and Basic Medical Science, Soochow University, Suzhou 215123, People's Republic of China.
authors.forename: Hao-Kun
authors.initials: HK
authors.lastname: Zhang
authors.forename: Guang-Li
authors.initials: GL
authors.lastname: Cao
authors.forename: Xiao-Rong
authors.initials: XR
authors.lastname: Zhang
authors.forename: Xiao-Juan
authors.initials: XJ
authors.lastname: Wang
authors.forename: Ren-Yu
authors.initials: RY
authors.lastname: Xue
authors.forename: Cheng-Liang
authors.initials: CL
authors.lastname: Gong
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000254:growth & development* / Q000378:metabolism; D012831:Bombyx / Q000254:growth & development* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D011679:Pupa / Q000254:growth & development* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D012604:Scorpion Venoms / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics
chemical_list: D004268:DNA-Binding Proteins; C052130:GAL4 protein, S cerevisiae; C445369:KK toxin, Buthus martensi; D029701:Saccharomyces cerevisiae Proteins; D012604:Scorpion Venoms; D014157:Transcription Factors; D049452:Green Fluorescent Proteins