Method for improving transgenic insect cell expression exogenous gene level

Abstract

The invention discloses a method for improving expression of extraneous genes by insect transgene cells. In the method, active promoter controlled neomycin resistance gene expression cassettes, enhancer elements and extraneous gene expression cassettes inside the cells of insects are cloned into a vector with reporter genes based on transposon piggyBAC factors; subsequently, the vector is mixed with subsidiary plasmids expressing transposase to transfect insect cell lines; transgene insect cells are acquired through sectionalized screening of G418. Engineering cells expressing extraneous genes at high level are acquired by cell clone technology in combination practical examination of expression level of extraneous genes. With the method, transgene insect cells can express extraneous genes at high level continuously; the expression products are free of rhabdoviruses with good bio-safety, and are processed perfectly as well as natural.

Description

technical field [0001] The invention relates to a transgene expression system, in particular to a method for increasing the expression level of exogenous genes in transgenic insect cells. Background technique [0002] There are two main insect baculovirus expression systems currently in use, one uses the alfalfa geometrid spodoptera nuclear polyhedrosis virus as a vector, and uses Fall Armyworm cells, cabbage caterpillar cells or insects as hosts; the other uses the silkworm nuclear polyhedrosis virus as a host. The body virus is used as the carrier, and the silkworm cell or silkworm worm and pupa are used as the expression host. Both expression systems use strong promoters such as polyhedrin gene or P10 gene or very early gene IE-1 of their respective viral vectors as regulatory elements for exogenous gene expression, by placing the exogenous target gene under the control of a strong promoter When the recombinant virus infects the host cell, a large amount of recombinant b...

AI Technical Summary

Problems solved by technology

Jarvis et al. transfected sf-9 cells with a plasmid carrying the neo marker gene and the ie-1 (very early gene of the californica nuclear polyhedrosis virus) promoter to control the expression cassette of the β-galactosidase gene, and passed the G418 Pressure screening to obtain a pure line of transformed sf-9 cells, the cells after 50 passages (about 6 months) still retain the integrated plasmid sequence, and the cells after 100 passages can still express homogeneous β-galactosidase, but The expression level is low, about 1% of the baculovirus expression vector system (BEVS) (J Cell Biochem, 1990, 42: 181-191)