44 results about "LacZ Genes" patented technology

The invention belongs to the technical field of biology, and mainly relates to a recombinant fowlpox virus transfer vector expressing a fowl adenovirus serotype-4 (FADV4) fiber2 gene, a constructing method thereof and applications of the transfer vector. A pMD19T-Simple vector is adopted as a base of the transfer vector. The FADV4 fiber2 gene, a lacz gene, and fowlpox virus genome replicated non-essential fragments which are LTYB and RTYB used for homologous recombination are inserted in a TA cloning site. The method includes constructing a plasmid pMD-TYB; constructing a plasmid pMD22; constructing a plasmid pMD22-lacz; constructing an intermediate vector pMD22-TYB-lacz; amplifying the FADV4 fiber2 gene; constructing a recombinant fowlpox virus transfer vector pMD22-TYB-lacz-F4; subjecting a chicken embryo Fibroblast to cotransfection with the transfer vector pMD22-TYB-lacz-F4 and a fowlpox virus; performing identification to select a positive product; performing subculture continuously; and identifying expression effects of the recombinant fowlpox virus. The recombinant fowlpox virus transfer vector constructed by the method lays a foundation for development of an efficient recombinant fowlpox virus genetic engineering living-vector vaccine expressing the fowl adenovirus serotype-4.

The invention discloses a pC series plasmid as well as a construction method and application thereof, wherein the pC series plasmid comprises a rep gene, a rop gene, a lacZ gene and an antibiotics resistance gene which are sequentially connected with one another end to end. The pC series plasmid has the characteristics of small molecular weight, large capacity, low copy number, high stability andthe like, can be used for instable genes with complicated structures and repeated sequence, cloning, screening and sequencing long section of genes, genome construction and other gene engineering fields.

The invention discloses a pC series plasmid as well as a construction method and application thereof, wherein the pC series plasmid comprises a rep gene, a rop gene, a lacZ gene and an antibiotics resistance gene which are sequentially connected with one another end to end. The pC series plasmid has the characteristics of small molecular weight, large capacity, low copy number, high stability andthe like, can be used for instable genes with complicated structures and repeated sequence, cloning, screening and sequencing long section of genes, genome construction and other gene engineering fields.

The invention relates to a mutant strain of the virulence gene VIR of aphthous ulcer virus and its preparation method and application. The preparation method comprises the method of cloning the gene fragment containing the VIR virulence gene of ORFV-SHZ1 strain and the flanking sequence by using the PCR method, and deleting it by the enzymatic cutting method. Remove the VIR virulence gene, insert the expression cassette with the late promoter P11 of vaccinia virus and the LacZ reporter gene at the deletion site, and construct a recombinant shuttle vector with the LacZ gene as a selection marker; transfect the recombinant shuttle vector into ORFV-infected Vero Homologous recombination was carried out in the cells, and the recombinant virus ORFV-ΔVIR-LacZ with deletion of the VIR virulence gene was obtained by plaque screening and purification. The invention quickly reduces the virulence of ORFV by means of genetic engineering, provides technical support for the research and development of ORF vaccines, has the characteristics of short production cycle, no need for domestication, convenient and quick operation, etc., and can be widely used.

The invention belongs to the field of biotechnology, and mainly relates to a recombinant fowlpox virus transfer vector expressing the fiber2 gene of chicken type 4 adenovirus (FADV4) and its construction method and application. The vector is based on the pMD19T-Simple vector, and the TA cloning site is inserted into the FADV4fiber2 gene, the lacz gene, and non-essential fragments LTYB and RTYB for genome replication of fowlpox virus used for homologous recombination. The method comprises the following steps: plasmid pMD-TYB; constructing plasmid pMD22; constructing plasmid pMD22-lacz; constructing intermediate vector pMD22-TYB-lacz; amplifying FADV4 fiber2 gene; constructing recombinant fowlpox virus transfer vector pMD22-TYB-lacz- F4: co-transfect chicken embryo fibroblasts with the transfer vector pMD22-TYB-lacz-F4 and fowlpox virus, and identify positive ones, and continue to subculture; identify the expression effect of the recombinant fowlpox virus. The recombinant fowlpox virus transfer vector constructed by the invention lays the foundation for the development of highly efficient recombinant fowlpox virus gene engineering live vector vaccine expressing chicken type 4 adenovirus.

The present invention relates to a non-toxic dipolar solvent for chromogenic substrate for detecting presence of lacZ gene and / or gene activity, which comprises a stabilizing amount of a solubilizing agent. The present invention also relates to a method for inducing lac operon in screening assay, comprising the step of contacting an agar plate with at least one essential oil in a concentration sufficient to induce the lac operon. The present invention further relates to a method for detecting the presence of bacteria, comprising the step of contacting an agar plate with at least one essential oil in a concentration sufficient to induce detection of the bacteria.

The present invention provides a fowlpox virus double-gene expression vector, which has the nucleotide sequence shown in SEQ ID NO.1, including promoter P7.5, LacZ gene, multiple cloning site on the LacZ gene, and rare enzyme Cutting site Not1, fowlpox virus DNA homology arms F11L1 and F11L2, AmpR resistance marker gene, origin of replication (ori). The dual-gene expression vector constructed by the present invention minimizes the length of the vector so that larger foreign genes can be accommodated; the number of available restriction sites is increased to facilitate the cloning of different foreign genes; the insertion site Not1 The promoter is not limited, and the promoter that matches with the exogenous gene can be freely selected, thereby improving the expression efficiency, and the expression efficiency of the exogenous gene can be improved by adopting the method of trans expression.

The invention provides a rapid screening system for Lactococcus lactis with knocked-in exogenous genes. The screening system comprises temperature-sensitive plasmids and the Lactococcus lactis, wherein the temperature-sensitive plasmids contain his gene segments and temperature-sensitive replicon-erythromycin resistance gene (Ts-Emr) segments; chromosomes of the Lactococcus lactis contain lacZ gene expression segments. lacZ genes of the Lactococcus lactis cannot be expressed when the exogenous genes are knocked in successfully, so that white colonies are shown on a solid medium containing X-Gal, otherwise, blue colonies are shown, the change from the blue colonies to the white colonies can be directly observed on the solid medium, the screening workload can be greatly reduced, and screening time can be greatly shortened.

The invention relates to a method for carrying out point mutations on essential genes on escherichia coli genomes by using the co-transformation of oligonucleotide and eliminable plasmids. Specifically, front four basic groups express recombinase in escherichia coli by using the co-transformation of oligonucleotide connected by sulfur phosphoryl, with a length of 91 basic groups, and having a mutation site in the middle, and eliminable plasmids. The eliminable plasmids contain homologous sequences of 50 bp lacZ genes, ampicillin resistance genes, I-SceI genes, two I-SceI cleavage sites and R6K replicators. In strains which are integrated with a 50 bp lacZ homologous fragment of a genome due to the catalysis of recombinase so as to express the amicillin resistance, the strains with point mutations are screened. Integrated strains can be induced to express I-SceI under the action of chlortetracycline subjected to heat inactivation, the I-SceI acts on cleavage sites thereof, and through the homologous recombination of two 50 bp lacZ fragments, the eliminable plasmids are eliminated.

The invention belongs to the technical field of biology, and mainly relates to a recombinant fowlpox virus transfer vector expressing a duck adenovirus serotype-2 (DADV2) fiber2 gene, a constructing method thereof and applications of the transfer vector. According to the transfer vector, a DADV2 fiber2 gene promoted by a fowlpox-virus-containing early-late promoter LP2EP2, a lacz gene promoted by a P11 promoter, and fowlpox virus genome replicated non-essential fragments which are LTYB and RTYB used for homologous recombination are inserted in a TA cloning site of a pMD19T-Simple vector. The method includes constructing a plasmid pMD-TYB; constructing a plasmid pMD22; constructing a plasmid pMD22-lacz; constructing an intermediate vector pMD22-TYB-lacz; constructing the transfer vector pMD22-TYB-lacz-DFB2; and subjecting the transfer vector pMD22-TYB-lacz-DFB2 to effect verification. The recombinant fowlpox virus transfer vector constructed by the method lays a foundation for development of an efficient recombinant fowlpox virus genetic engineering living-vector vaccine expressing the duck adenovirus serotype-2.