Quantitative analysis of cytoplasmic actin gene promoter and nuclear polyhedrosis virus immediate-early promoter activities in various tissues of silkworm Bombyx mori using recombinant Autographa californica nuclear polyhedrosis virus as vector.
Cassettes harboring luciferase reporter driven by Bombyx mori cytoplasmic actin gene promoter (A3) (671 bp) and B. mori nuclear polyhedrosis virus immediate-early promoter (IE-1) (580 bp) were transferred to the bacmid AcDeltaEGT to generate the recombinant Autographa californica nuclear polyhedrosis viruses, AcNPVA3Luc and AcNPVIELuc, respectively. Recombinant baculoviruses were injected into the hemocoele of newly ecdysed 5th instar larvae. The activities of the A3 and IE-1 promoters in various tissues were measured by luciferase activity assay and normalized by the copy number of recombinant virus. Results showed that the activity of the A3 promoter was approximately 10-fold higher than the IE-1 promoter. The promoter activities of A3 and IE-1 were highest in the silk gland, followed by fat body, middle gut, malpighian tubule, and hemocyte. In silk gland, activity of the two promoters was highest in posterior silk gland, followed by middle and anterior silk glands. The difference in promoter activities reflects the growth speed of tissue in silkworm larvae. The activity of the A3 promoter remained unchanged and was not inhibited significantly by viral factors at least 3-4 d post injection of rAcNPV.
School of Pharmaceutical Engineering, Shenyang Pharmaceutical University, Shenyang 110016, China.
Yi
Y
Zhang
Xue
X
Zhang
Huanzhang
H
Xia
Yuegui
Y
Xue
Jianyang
J
Wang
Baozhong
B
Tian
Zhenguo
Z
Wei
Changde
C
Lu
D000199:Actins / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D012831:Bombyx / Q000502:physiology*; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D024861:Genetic Enhancement / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics*; D017924:Nucleopolyhedroviruses / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics; D014162:Transfection / Q000379:methods*
D000199:Actins; D011994:Recombinant Proteins