Method for constructing recombinant expression vector simultaneously expressing a plurality of genes

Abstract

The invention discloses a method for constructing a recombinant expression vector simultaneously expressing a plurality of genes. An MISSA system consists of a strain and a vector, wherein the strain comprises a donor strain and an acceptor strain; the vector comprises a donor vector and an acceptor vector; replication initiation protein carried by a chromosome of the donor strain is responsible for replicating and amplifying a suicidal donor vector, and the carried conjugal transfer protein Tra is responsible for conjugal transfer of the donor vector; the acceptor strain can induce and express two sets of locus specific recombinant proteins, namely Cre recombinant enzyme and lambda phage locus specific recombinant proteins; when the donor strain is mixed with the acceptor strain, the donor strain and the acceptor strain are conjugated, and the donor strain is transferred into the acceptor vector because the donor vector in the donor strain carries an oric element; and the donor vector and the acceptor vector undergo recombination reaction in the acceptor strain under the action of the two sets of locus specific recombinant proteins, so that a target gene carried by the donor vector is assembled onto the acceptor vector. The process is repeated, so a plurality of target genes can be assembled onto the acceptor vector according to the preset direction and order.

Description

technical field

[0001] The invention relates to a method for constructing a recombinant expression vector for simultaneously expressing multiple genes. Background technique

[0002] Many agronomic traits, including crop yield traits, metabolic pathways, protein complexes and signal transduction pathways, are controlled by polygenes. Genetic manipulation of multiple genes has important application prospects for both basic research and bioengineering. Golden rice (Ye X, Al Babili S, Kloti A, Zhang J, Lucca P, Beyer P, Potrykus I (2000) Engineering the provitamin A (beta-carotene) biosynthetic pathway into (carotenoid-free) riceendosperm. Science 287: 303- 305) and purple tomato (Butelli E, Titta L, Giorgio M, Mock HP, Matros A, Peterek S, Schijlen EG, Hall RD, Bovy AG, Luo J, Martin C (2008) Enrichment of tomato fruit with health-promoting anthocyanins by expression of select transcription factors.Nat Biotechnol 26:1301-1308) are two successful cases of multi-gene transgenic...

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