41 results about "Polyadenylic acid" patented technology

The invention relates to a scallop protein H2A gene clone and the monoclonal expression technique of N end antibiotic polypeptide in the field of molecular biology technique. It uses isogenesis clone technique to obtain the scallop group protein H2A gene group total length; the gene DNA has a 375bp open reading frame with the coding 125 amino acids, 3' non-coding area comprises two ending signals which are stem ring structure and polyadenylic acid rear signal. It uses pPIC9K expressing carrier to express the protein N end 30aa in Pichia pastoris, the expressing product has bacterial inhibition to the Micrococcus luteus and the Vibrio splendidus.

The invention discloses a silkworm PABP (polyadenylic acid bonding protein) bonding protein interacting factor gene BmPaip1. The nucleotide sequence of the gene BmPaip1 is shown as SEQIDNO.3, the open reading frame of the gene BmPaip1 is 1194bp long, composed of eight exons and codes 398 amino acids, theoretical protein molecular weight is 44kDa, and isoelectric point pI is equal to 5.02; and SMART online software analysis shows that the bits from 100 to 398 in the amino acid sequence of the gene BmPaip1 of a silkworm are highly homologous with an eIF4G protein of an eucaryon, and expression detection in insect cells shows that expression level of a reporter gene LUC (luciferase) can be obviously improved in cells genetically modified by the gene BmPaip1 of the silkworm, so that the gene BmPaip1 of the silkworm can be combined with eIF4E and eIF4A to form a translation initiation complex and can be used for improving translation efficiency of an exogenous gene.

The invention discloses an eucaryon expressing shuttle vector as well as a constructing method and applications thereof. The eucaryon expressing shuttle vector adopts an annular vector which comprises a prokaryon replication origin, two eucaryon replication origins, selective marker genes and a foreign gene expressing box; the foreign gene expressing box is composed of two promoters sharing an identical duplicating direction, connecting segments and polyadenylic acid tailing signals from upstream to downstream sequentially; and the connecting segment contains an EcoRV recognition sequence. The process of constructing the recombinant vector of expression foreign gene through the eucaryon expressing shuttle vector is convenient, quick, economical and highly efficient, and the vector contains two eucaryon replication origins, therefore, the vector can be greatly appreciated in cells expressing T antigen, and the protein expressing level can be improved. The eucaryon expressing shuttle vector has broad application prospect.

The invention discloses an expression vector expressing green fluorescent protein by fusion and a construction method and application thereof. The vector is in ring shape , comprising a procaryon replication origin, two eucaryon replication origins and selective marker genes, and two green fluorescent protein gene expression cassettes; the green fluorescent protein gene expression cassettes, fromupstream to downstream, consist of two promoters with the same transcription direction, junction fragments, green fluorescent protein encoding genes and polyadenylic acid AATAAA; the green fluorescent protein encoding gene lacks an initiation codon. The junction fragment comprises an EcoRV recognition sequence; the first place and the second place of the green fluorescent protein encoding gene from the end of 5' overlaps the last two places from the end of 5' of the EcoRV recognition sequence; the junction fragment comprises or does not comprise the initiation codon; when the junction fragment comprises the initiation codon, the initiation codon has a space of 2+3n or 1+3n from the first place from the end of 5' of the EcoRV recognition sequence, wherein, n is a positive integer. The vector comprises a green fluorescent protein gene, and can only express the green fluorescent protein on the condition that the gene is correctly expressed, therefore, the interference due to the expression of fluorescence by an initial vector can be effectively excluded.

The present invention provides methods for stably integrating and / or expressing one or more recombinant polynucleotides in a host cell. The recombinant polynucleotide is usually integrated into the host genome at some natural chromosomal integration site. The integration can be mediated by homologous recombination or with hybrid recombinases targeted to specific chromosomal locations. Stable integration and strong transcriptional activity of foreign genes are supported by native chromosomal integration sites in the host cell, which are located within or near specific genes in the CHO genome, the ankyrin 2 gene (Ank2), cleavage and polyadenylation-specific factor 4 gene (Cpsf4), C-Mos gene and Nephrocystin-1 / Mal gene. Also provided are methods and nucleic acid molecules for the insertion of site-specific recombination sequences (chromosomal landing pads) into these specific chromosomal locations. Further provided are engineered host cells containing chromosomal landing pads or transgenes integrated into natural chromosomal integration sites. Further provided are methods and compositions (eg, kits) for integrating and expressing heterologous polynucleotides in host cells with inserted chromosomal landing pads.