1522 results about "Molecular Biology Technique" patented technology

The present invention provides methods for constructing peptide construct sets and methods of use of these peptide construct sets in assay systems for peptide analysis, and in particular for use in high throughput peptide analysis. The methods allow for analysis of large sets of peptide constructs in a cost-effective manner, employing molecular biological techniques that are both robust and easily parallelized. Thus, the methods allow for the construction of peptide construct sets encompassing, e.g., the human proteome.

The present invention discloses a tobacco cultivation method. Said method is characterized by that it adopts a molecular biological technique to identify key gene capable of promoting axillary bud germination and growth after the tip is pruned, then uses nocuity inducible promoter to drive antisense RNA and make it promptly express after the tip is pruned so as to inhibit the expression of said key gene capable of promoting axillary bud germination and growth and make the axillary bud can not be germinated or grown.

The invention relates to the microbiology and molecular biology technical field, in particular to a method for extracting pig manure sample total DNA. The method mainly comprises manure sample pretreatment and sample total DNA extraction. Moreover, the method comprises the following steps that: collected fresh manure sample or frozen manure sample is taken as a material; the pretreatment of the manure sample is carried out by means of a PBS buffer solution, a paraformaldehyde solution and anhydrous alcohol so as to obtain manure sample bacteria suspension; and a CTAB solution and beta-mercaptoethanol, etc. are used to carry out cell fragmentation, thereby releasing DNA. The method has the characteristics of simple operation, quickness, high efficiency and ideal repeatability and economical efficiency, etc.; moreover, the extracted DNA has high concentration, purity and yield, and can be directly used in subsequent molecular biology experimental researches such as PCR amplification without purification.

The invention belongs to the fields of molecular biotechnology and experimental methods and specifically relates to a fluorescence quantitative PCR reaction solution. The solution is characterized by comprising a PCR reaction reinforcing agent which is one or more selected from the group consisting of dimethyl sulfoxide, glycerin, bovine serum albumin, methanamide, polyethylene glycol 6000, spermidine, ammonium sulfate and betaine, e.g., dimethyl sulphoxide, bovine serum albumin and betaine. The fluorescence quantitative PCR reaction solution further comprises heatproof DNA polymerase having 5'-3' polymerase activity and 5'-3' exonuclease activity and preferably comprises heatproof DNA polymerase and hot-start heatproof DNA polymerase, both of which having 3'-5' exonuclease activity. The invention also relates to a kit containing the reaction solution and a method of applying the reaction solution in fluorescence quantitative PCR.

The present invention relates to a bovine VDJ cassette (BF1H1) that provides the novel ability to develop chimeric immunoglobulin molecule capable of incorporating both linear T cell epitope(s) (CDR1H and CDR2H) as well as conformational B cell epitope(s) (exceptionally long CDR3H). Further, multiple epitopes can be incorporated for development of multivalent vaccine by replacing at least a portion of an immunoglobulin molecule with the desired epitope such that functional ability of both epitope(s) and parent VDJ rearrangement is retained. The antigenized immunoglobulin incorporating both T and B epitopes of interest is especially useful for development of oral vaccines for use in humans apart from other species including cattle. The long CDR3H in BF1H1 VDJ rearrangement originates from long germline D-genes. The novel bovine germline D-genes provide unique molecular genetic marker for sustaining the D-gene pool in cattle essential for immunocompetence via selective breeding. D-gene specific DNA probe permits typing and selection of breeding cattle stock for maximum gemline D gene pool for better health and disease prevention. The bovine D-genes are unique to cattle and, therefore, provide sensitive and specific forensic analytical tool using molecular biology techniques to determine tissues suspected of bovine origin.

The invention discloses an omega-aminotransferase mutant gene and an application thereof, and belongs to the field of the molecular biological technology. The nucleotide sequence of the omega-aminotransferase mutant gene is shown as SEQ ID NO.1. The omega-aminotransferase amino acid sequence embodied by the code of the omega-aminotransferase mutant gene is shown as SEQ ID NO.2. The omega-aminotransferase mutant gene is firstly provided, and the omega- aminotransferase corresponding to the code of the omega-aminotransferase mutant gene has a high enzyme amount, so that the biological catalysis and conversion efficiency is improved in industrial production. The invention further provides the application of the omega-aminotransferase embodied by the code of the omega-aminotransferase mutant gene in preparation of trifluoro-propylamine. The trifluoro-propylamine is added to a buffer solution to serve as the substrate for the reaction, and then products of trifluoromethyl amine compounds are collected from the reaction liquid. The method for preparing the trifluoro-propylamine has the obvious advantages that the reaction conditions are mild, no pollution is caused, and the process is simple.

The invention relates to lipase mutants with enhanced thermal stability, which belongs to the technical field of enzyme gene engineering. Rhizopus chinensis CCTCC No.M201021 lipase used as the parent is processed by molecular biotechnology to obtain the lipase mutants with enhanced thermal stability. In the amino acid sequences of the mutants, the related amino acid mutation(s) is(are) one or a plurality of Met101Thr, Glu107Gly, Ala129Ser, Ser151Asn, Cys160Leu, Lys161Arg, Pro168Leu, Pro168His, Leu180His, Asp182Tyr, Thr183Ala, Thr218Ser, Lys219Asp, Ala230Phe, Ser234Phe, Val261Gly, His317Pro, Val329Ala, Glu363Arg, Asn366Asp and Ser373Cys. The mutants are expressed by half-life period t50 at 65 DEG C. The thermal stability of the mutants is enhanced as compared with the parent Rhizopus chinensis lipase. The invention also discloses a DNA sequence, an expression carrier and a host cell for coding the lipase mutants.

The invention relates to a quick, sensitive and high-throughput intestines source pathogenic bacterium detection method. The detection method provided by the invention integrates two powerful molecular biological techniques: polymerase chain reaction PCR and a micro-array, and directly fixes a probe of PCR hybridization in a hybridization cabin of the micro-array on a same chip with a PCR reaction chamber. The detection method comprises the following steps of enriching bacteria; extracting a DNA solution; carrying out PCR amplification; hybridizing; cleaning; and judging the result. The method provided by the invention can quickly detect genes of vibrio parahaemolyticus, Shigella, staphylococcus aureus, listeria monocytogenes and salmonella in high throughput, and the detection efficiency of front-line inspection and quarantine personnel of import and export ports can be greatly improved, thereby not only reducing the workload, but also solving the undetected positive result problem probably caused by conventional detection method to the maximum extent. Therefore, food safety incidents are prevented to the maximum extent.

An Escherichia coli recombinant strain producing shikimic acid, and a construction method and application thereof belong to the technical field of microbial gene engineering. The invention firstly utilizes a molecular biology technique to delete shikimic acid kinase I gene (aroK) and shikimic acid kinase II gene (aroL) of Escherichia coli CICIMB0013, and a gene (ptsG) of a key protein EIIBC<Glc> and a quinin acid/shikimic acid dehydrogenase gene (ydiB) of a glucose phosphotransferase system to obtain an Escherichia coli mutant strain CICIMB0013.SA4 (delta aroK, delta aroL, delta ptsG, delta ydiB). The invention also constructs a recombinant expression plasmid pTHGAA containing key genes comprising aroG*,ppsA and tktA in a metabolic pathway of shikimic acid; and the recombinant expression plasmid pTHGAA is transferred into the recombinant strain CICIMB0013.SA4 to obtain a recombinant Escherichia coli B0013 (SA4/pTHGAA) capable of producing shikimic acid efficiently. The Escherichia coli recombinant strain provided by the invention can realize efficient accumulation of shikimic acid in a fermentation process.

The invention belongs to the technical field of molecular biology and provides primers, a probe composition and a kit for rapid identification of nine animal origin ingredients in food or feed, a detection method for identification of the nine animal origin ingredients in the food or feed and application of the primers, the probe composition, the kit and the detection method. Origin ingredients of the food or feed containing multiple species can be identified rapidly by the primers, the probe composition and the real-time fluorescent PCR (polymerase chain reaction) joint detection kit. The detection method includes: designing a universal primer by taking 16SrDNA as a target gene, and designing specific probes of nine species by designing to construct internal amplification control and designing the specific probes aiming at internal control sequences; performing PCR amplification by three PCR reaction systems; interpreting the origin ingredients of the nine species directly through corresponding fluorescent probe signals and Ct values. The detection method is low in cost, time saving and high in efficiency and can achieve identification of multiple species simultaneously.

The present invention is generally directed to novel functionalized biomolecules and methods for generating such biomolecules. Biomolecules may generally include nucleic acids, peptides, multicomponent molecular complexes and/or any other molecular products that may be produced by living organisms. The present invention is further directed to cells and/or organisms manipulated to produce such functionalized biomolecules. The cells contemplated by the present invention include both prokaryotic as well as eukaryotic cells. The functionalized biomolecules are produced via materials introduced into the cell using standard molecular biology techniques or are incorporated within the genomic nucleic acid of a cell by standard recombination techniques. Further contemplated is the use of such cells for sequestration of target molecules within the cells.

The invention relates to a bacillus licheniformis engineering bacterium for efficiently synthetizing poly-gamma-glutamic acid, and belongs to the field of microbial metabolism engineering. A conventional molecular biological technique is used; on the basis of bacillus licheniformis WX-02 stored in a laboratory, a double-ingredient regulation and control factor degU is expressed in an enhanced way through external sources; the bacillus licheniformis engineering bacterium WX-02/pHY-degU capable of efficiently synthetizing the poly-gamma-glutamic acid under the condition of no addition of an external source of glutamic acid is obtained. Liquid fermentation proves that the concentration of the poly-gamma-glutamic acid at a fermentation end point reaches 32.78 g/L; compared with that of an original bacterial strain of bacillus licheniformis WX 02, the concentration is improved by 47.59 percent; the synthesis capability of the poly-gamma-glutamic acid is obviously enhanced. The bacterial strain is preserved in CCTCC (China Center For Type Culture Collection) on May 27, 2016, and the preservation number is CCTCC NO:M2016294.

The invention relates to a method for studying the structural diversity of a daqu bacterial community, i.e. the denaturing gradient gel electrophoresis (DGGE) technology, which belongs to the technical field of microbial ecology and mainly comprises the following steps that: 1), daqu genomic deoxyribonucleic acid (DNA) is directly extracted; 2), bacterial universal primers are selected, and specific DNA fragments in the bacterial ribosome DNA are amplified; 3), polymerase chain reaction (PCR) products are separated through DGGE; 4), corresponding strips of microbes in the DGGE fingerprint map are recovered in a gel cutting way; 5), PCR is carried out again, products are connected to a T carrier, the blue and white spot screening is carried out, and positive clone verification is carried out; and 6), the species information of the corresponding microbes of the DGGE strips is obtained through the sequence test. The method does not rely on the molecular biology technology of the traditional culture method, has the characteristics of sensitivity, convenience and accuracy and solves the difficult problems to study some microbes incapable of being cultured or difficult to culture, and the method provides the technical basis for the verification of the daqu bacterial community structure and the discovery of new wine brewing microbes or functional microbes.

The invention belongs to the molecular biology technology field, and concretely relates to a method for sequencing and development of Asplenium nidus L. EST-SSR primers based on transcriptome. The method comprises the following steps: firstly, a transcriptome library is constructed; secondly, transcriptome data is obtained, namely, sequencing data is subjected to splicing by utilization of a Trinity software and sequences are spliced into a complete transcriptome; thirdly, SSR site search is carried out, namely, combined with a Perl programming language method, a lot of sequence information is subjected to batch processing, and SSR site search is carried out; fourthly, batch design of EST-SSR primers is carried out. Different fiddlehead materials are selected to verify the designed SSR primers, if strips are detected, successful SSR primers are obtained. Through the method, 4063 pairs of SSR primers are designed, and a new method and thought are provided for development of Asplenium nidus L. EST-SSR primers and further for achieving molecule label auxiliary selection breeding.

The invention belongs to the technical field of molecular biology, and particularly relates to a kit for extracting genome DNA (Deoxyribose Nucleic Acid) from plant leaves based on a paramagnetic particle method and an extracting method thereof. The kit for extracting genome DNA from plant leaves based on the paramagnetic particle method comprises a pretreatment solution, a rapid cracking liquid, a DNA binding liquid, a magnetic bead suspension and an eluent, wherein cell walls and cell membranes can be effectively cracked through the pretreatment solution; and the interference of impurities such as polysaccharides, polyphenol, tannin and the like on genome DAN extraction can be effectively eliminated through the pretreatment solution. In the rapid cracking liquid, guanidine hydrochloride and Tween 20 are taken as main components, so that plant cells can be fully cracked at the normal temperature within 2 minutes, only two minutes are required in a subsequent centrifuging step, and the operation time is greatly saved. The guanidine hydrochloride is a strong denaturant, has a good cell cracking effect. Moreover, organic solvents such as chloroform and the like are not required to be added, so that damages to operating personnel are avoided, and safety and reliability are achieved.

The invention relates to miRNAs (micro ribonucleic acids) used for detecting prostatic cancer and a kit, and belongs to the technical field of molecular biology, wherein the miRNAs comprise carcinogenic miRNAs and cancer-inhibiting miRNAs; expression quantity of the carcinogenic miRNAs in real-time PCR (polymerase chain reaction) method detection, and expression quantity of the cancer-inhibiting miRNAs in the real-time PCR method detection are lowered. According to different degrees of the prostatic cancer, different carcinogenic miRNAs and cancer-inhibiting miRNAs are adopted to respectively carry out diagnosis in a targeted manner. A technical scheme provided by the invention is adopted, so that diagnosis and prognosis prediction results of the prostatic cancer can be quickly, simply, efficiently and stably obtained.

The invention relates to a cloning vector and preparation and application thereof and discloses a cloning vector pUC57-ccdB which is a modified vector with ccdB genes inserted at multiple cloning sites of a pUC57 vector, wherein the ccdB genes have blunt-end restriction enzyme digested recognition sites. By means of the lethal effect of CcdB protein on escherichia coli not containing F plasmids, the ccdB genes having the restriction enzyme Sma I digested sites are inserted on the pUC57 plasmids through the molecular biological technology to obtain the vector pUC57-ccdB. Blunt ends are generated through Sma I digestion and are connected with the genes needing to be cloned so that insertion of the genes needing to be cloned can be achieved. Meanwhile, bacterial colonies with empty vectors are avoided. The cloning vector plays an important role in the fields of molecular biology and genetic engineering.

The recombination, expression and application of Chinese prawn's antibacterial peptide gene are disclosed. Its recombination expression method includes configuring recombinant carrier (pET series, pGEX-4T series, pPIC series, pAO815, and baculovirus expression carrier), transferring the carrier to colibacillus and yeast and transfecting insect cells, screening engineered bacterial strain or cell, and inducing expression. The expressed product can be used to prepare feed additive, antistaling agent, cosmetics and medical products.

The invention discloses a lipase mutant with increased optimum temperature, which belongs to the technical field of enzymatic genetic engineering. The lipase mutant with increased optimum temperature disclosed by the invention is obtained through taking rhizopus chinensis China center for type culture collection (CCTCC) M201021 lipase as a parent and utilizing a molecular biological technique, and the mutant amino acid of the mutant is Asp310Val. The optimum temperature of the lipase mutant is increased compare with the optimum temperature of the parent of rhizopus chinensis lipase and has an important industrial application value.