368 results about "Mutation rate" patented technology

Dominant negative alleles of human mismatch repair genes can be used to generate hypermutable cells and organisms. By introducing these genes into cells and transgenic animals, new cell lines and animal varieties with novel and useful properties can be prepared more efficiently than by relying on the natural rate of mutation. These methods are useful for generating genetic diversity within immunoglobulin genes directed against an antigen of interest to produce altered antibodies with enhanced biochemical activity. Moreover, these methods are useful for generating antibody-producing cells with increased level of antibody production. The invention also provides methods for increasing the effector function of monoclonal antibodies and monoclonal antibodies with increased effector function.

The invention discloses a CRISPR/Cas9 system-containing targeted knockout vector and adenovirus and applications thereof. The targeted knockout vector is prepared through the following steps: after a pX330 U6-Chimeric_BB-CBh-hSpCas9 plasmid is subjected to enzyme digestion and filling-in by using EcoRI and SacII, connecting the pX330 U6-Chimeric_BB-CBh-hSpCas9 plasmid with a pAdTrack-CMV plasmid subjected to enzyme digestion and filling-in by using BstXI; after the obtained product is linearized by using BbsI, connecting the obtained product to a specific target sequence of a desired gene; and after the obtained object is linearized by using PmeI and dephosphorylated by using CIAP alkaline phosphatase, recombining the obtained product with a pAdEasy-1 plasmid. The targeted knockout vector can mutate gene sequences in target sequence areas, and the mutation rate is high, up to 30.6-45.8%, therefore, the targeted knockout vector can be used for gene site-directed mutation, and lays a foundation for gene therapy.

A method of assigning web service requests to service providers includes searching for an optimal assignment from all possible assignments using a genetic algorithm (GA) that represents possible assignments as chromosomes, and converging towards an assignment of web service request to service providers that maximizes overall business value for all workflows to the service providers. An adaptive mutation scheme is used to introduce mutation into populations of chromosomes. The mutation scheme includes a mutation rate that increases when chromosomes under evaluation fail to improve its workload against the metric over a certain number of generations.

The invention relates to the field of microorganisms, in particular to a site-directed mutation method for genomes of saccharomyces cerevisiae. To-be-repaired single-base mutation sites on the genomes of the saccharomyces cerevisiae can be repaired by the aid of CRISPR/Cas9 technologies. Effects can be realized near the to-be-repaired sites by Cas9 proteins under the guidance actions of guide RNA (ribonucleic acid) if the mutation sites are positioned in PAM sequences (NGG) or front 11bp of the PAM sequences, and the genomes of the saccharomyces cerevisiae can be subjected to double-strand break at cut positions, so that donor DNA (deoxyribonucleic acid) can be efficiently recombined. Compared with existing results, the site-directed mutation method has the advantages that the sites of the genomes of the saccharomyces cerevisiae can be efficiently and quickly mutated, and screening markers do not need to be integrated for the genomes of the saccharomyces cerevisiae.

The invention discloses a primer and probe for detecting a human epidermal growth factor receptor (EGFR) gene and/or a K-ras gene, a kit containing the primer and the probe and a device for detecting genetic mutation on the basis of a digital PCR platform.The method for detecting the genetic mutation by means of the primer and the probe comprises the steps that the prime and the probe are provided; DNA of a sample to be detected is extracted; a fluorescent PCR reaction system capable of amplifying a mutant gene sequence is prepared; a target probe and an internal reference probe are utilized to be hybridized with amplified products respectively, and fluorescent signals of corresponding fluorescent groups are detected; existence of the genetic mutation is judged and/or the mutation rate is calculated according to the strength and proportion of the fluorescent signals of the target probe and the internal reference probe.According to the method for detecting the genetic mutation, the needed primers and probes are small in number, the optimization procedure is simple, related mutation of EGFR and/or K-ras gene can be qualitatively or quantitatively detected, and the detection sensitivity is high; a DNA sample with low initial amount can also be detected stably.

The invention relates to a method for reducing CRISPR/Cas9 mediated embryo gene editing missing rate. The method comprises the following steps: compounding a sgRNA sequence of a targeting gene and constructing a sgRNA expression plasmid carrying T7 promoter; in vitro transcribing Cas9mRNA and sgRNA; and importing intracytoplasmic sperm into unfertilized MII ovum by adopting intracytoplasmic sperminjection (ICSI) and meanwhile importing Cas9mRNA and sgRNA. According to the method provided by the invention, the targeting editing efficiency is high and the missing mutation rate is low.

The invention discloses a CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 recombinant lentiviral vector containing gRNA sequence specifically targeting CCR5 and application thereof. A lentivirus of the CRISPR/Cas9 recombinant lentiviral vector containing gRNA sequence specifically targeting CCR5 gene Delta 32 region is constructed that the lentivirus can introduce cells into a CRISPR/Cas9 system specific to CCR5, double-chain breakage occurs to a specific site of CCR5 gene, a random mutation is introduced to a breakage site after repairing by means of nonhomogeneous recombinant terminal binding, and the mutation rate reaches 90% and above. As gRNA is a nonhomogeneous region of CCR5 and CCR2, detection shows that the missing efficiency of the two gRNAs is lower than 0.2%. Cells modified via the recombinant lentivirus have significantly decreased efficiency of HIV (human immunodeficiency virus) infection. The system is quick to construct, simple and low in price, and is applicable to gene therapy of acquired immune deficiency syndrome.

The invention discloses a method and a device for route optimization of logistics delivery vehicle, and belongs to the technical field of logistics. The method comprises the following steps of: initializing a congestion matrix alpha and a distance matrix D, generating a delivery route weight matrix omega=alpha D, and initializing a population module N<ZQ>; selecting a population size N<X>, a maximum number of generations N<G>, a crossing-over rate beta, a mutation rate gamma and a number of generations n=0, generating an initial route r1 through a greedy algorithm, and performing mutation operation on the initial route r1 to generate N<ZQ>-1 new routes; calculating fitness A<n> of each route of a first generation population formed by the initial route and the new routes, selecting N<X> routes with the highest fitness from the current population by adopting selection operators, and performing crossover and mutation operations on the N<X> routes to generate a population of next generation; updating n=n+1, when n=N<G>, calculating the fitness A<n> of all the routes in the latest population, and selecting the delivery route with the highest fitness in the current population as the optimal route. According to the invention, when the logistics delivery vehicle delivers goods, the delivery time can be as less as possible, and the delivery route can be as short as possible.

The invention relates to an automatic calibration parameter optimization method of an engine based on genetic algorithm. The method comprises the following steps: introducing a genetic algorithm based on the original calibration technology; and establishing the corresponding original population, the fitness evaluation formula, the crossover rate, the mutation rate and the terminal conditions for each parameter by utilizing the genetic algorithm when the engine control parameters are optimized, and carrying out genetic algorithm optimization operation, thus the optimal gene is obtained as the control parameter in an MAP. The method has the beneficial effects that the method does not rely on any subjective factor, and has high objectivity, as long as engineers provide boundary conditions of the calibration objects and the final optimizing target, the optimization configuration of the selected parameters can be carried out automatically through the method; simultaneously the method has very high efficiency, multiple iterations can be carried out in very short time, thus the optimal solution can be obtained rapidly; and the method has high spreadability, and can spread the whole constrained range, thus the accuracy of optimization is improved.

The invention discloses a fluorescent labeling composite amplification kit for 30 STR (short tandem repeat) loci of human Y chromosome and application of the kit. It is possible to multiple amplify 30 low-mutation-rate STR loci on Y chromosome in single reaction; by designing locus arrangements and specific primer sequences, 30 loci are divided into 5 groups, and six-color fluorescent system marks are used; the Y-STR kit is the domestically first one employing six-color fluorescent detection technology and is an STR kit detecting a highest number of loci, all the loci included are low-mutation-rate Y-STR loci, and the kit is more suitable for examining male families; the composite amplification kit is good in primer specificity, can tolerate a wide range of temperatures and is highly adaptive to subject materials.

The invention relates to a method for preparing an antibody-producing cell line capable of directed constitutive hypermutation of a specific nucleic acid region, comprising the steps of: a) screening a clonal cell population for V gene diversity; b) isolating one or more cells which display V gene diversity and comparing the rate of accumulation of mutations in the V genes and other genes of the selected cells; and c) selecting a cell in which the rate of V gene mutation exceeds that of other gene mutation.

A molecule adaptor having characteristics of high stability, a high sample DNA connection efficiency and correction functions is designed based on illumina sequencing adaptor optimization. The molecule adaptor can detect mutation sites the mutation frequency of which is as low as 0.05%. The molecule adaptor is used for identifying real mutations in a sample sequencing library constructing process and false-positive mutations introduced by an operating process. In addition, a method of constructing a sequencing library for a sample to be detected is provided.

Dominant negative alleles of human mismatch repair genes can be used to generate hypermutable cells and organisms. By introducing these genes into cells and transgenic animals, new cell lines and animal varieties with novel and useful properties can be prepared more efficiently than by relying on the natural rate of mutation. These methods are useful for generating genetic diversity within immunoglobulin genes directed against an antigen of interest to produce altered antibodies with enhanced biochemical activity. Moreover, these methods are useful for generating antibody-producing cells with increased level of antibody production. The invention also provides methods for increasing the affinity of monoclonal antibodies and monoclonal antibodies with increased affinity.

The invention provides an exome potential pathogenic mutation detection method based on a family line. The detection method comprises the following steps: 1) reading a result file of an exome sequencing data processing flow, and conducting function filtering; 2) reading the file obtained in the last step, extracting mutations in all samples, calculating a union set, and then combining all samples, so that a matrix is constituted; 3) extracting mutation information in the matrix obtained in the last step, enumerating and assessing pathogenicity of single mutation and pathogenicity of combined dual-site mutation, so that a potential pathogenic mutation list is obtained; and 4) in accordance with the list obtained in the last step, calculating the appearance situations of sites in various samples and target genes. According to the method disclosed by the invention, data integration and basic filtration are completed by taking an output result of the common exome sequencing processing flow as an input condition; by virtue of a special mutation screening algorithm, a candidate set of the potential pathogenic mutations is provided; and the method focuses on solving a problem on potential pathogenic mutation mining of sequencing data with high heterogeneity, high mutation rate and high noise.

The invention discloses application of Y-STR in individual identification. The Y-STR is selected from any one of or a combination of over any two of DYG1-DYG36. The invention also discloses a detection kit and a detection method used for individual identification. The invention applies 36 new Y-STR loci with high mutation rates effectively in individual identification, and the resolution and accuracy are substantially raised.

The invention discloses a fluorescence labeling composite amplification kit for human Y chromosome 27 STR gene loci, which can simultaneously amplify 27 Y chromosome STR gene loci (including 24 low-mutation-rate Y chromosome STR gene loci and 3 high-mutation-rate Y chromosome STR gene loci) in a single multiplex reaction. By designing the unique gene locus configuration mode and specific primer sequences, the 27 gene loci are divided into 4 groups which are used for different fluorescence labelings. The composite amplification system has the advantages of wide temperature resistance range, high adaptability of the detection material, and high identification capacity for irrelevant and close relative male individuals.

A system for monitoring municipal saprobia inhibitive ability in real time based on oxygen uptake rate (OUR) comprises an aeration siphon device communicated with a sewage supplying system and a sludge supplying system. The aeration siphon device is composed of an aeration groove and a siphon groove in adjoining mode, the wall of the aeration groove is higher than that of the siphon groove, the aeration groove is communicated with the sewage supplying system and the sludge supplying system, and the siphon groove is communicated with an inlet of a closed coiler flow-pushing type bioreactor through a siphon tube. A dissolved oxygen (DO) detection system detects DO values of the inlet and an outlet of the closed coiler flow-pushing type bioreactor, and the signal output end of the DO detection system is connected onto a programmable logic controller (PLC) control system which is connected with an alarm system. The core part of system for monitoring municipal saprobia inhibitive ability in real time adopts the closed coiler flow-pushing type bioreactor so as to monitor the DO value of the inlet and the DO of the outlet in real time, automatically calculate mutation condition of activated sludge OUR, determines OUR mutation rate with different inhibition degree through plenty of experiments, and judge inhibitable degree of the saprobia by comparing automatically calculated OUR mutation rate with a preset value.

The invention discloses oriented domestication and breeding of methylotrophic bacterium capable of producing pyrroloquinoline quinone at high yield. The invention provides an oriented domestication and breeding method for preparing a methylotrophic bacterium mutant strain capable of producing pyrroloquinoline quinone at high yield. The method comprises the following steps: 1) carrying out repeated induced culture on an initial methylotrophic bacterium used as a treatment object in a methanol-containing liquid culture medium to obtain the domesticated bacterium, wherein all the domesticated bacteria constitute a domesticated strain library; 2) fermenting the domesticated bacteria in the domesticated strain library, and collecting all the domesticated bacterium fermentation liquids; and 3) detecting pyrroloquinoline quinone in all the domesticated bacterium fermentation liquids, and selecting the single strain, the fermentation liquid of which has higher pyrroloquinoline quinone yield than the initial methylotrophic bacterium fermentation liquid, thereby obtaining the methylotrophic bacterium mutant strain capable of producing pyrroloquinoline quinone at high yield. The methanol concentration in the culture medium is gradually enhanced to perform stress domestication on the methylotrophic bacterium, thereby enhancing the breeding direct mutation rate and the pyrroloquinoline quinone yield increase amplitude.

The invention relates to a method for creating a new peanut specie, belonging to the technical field of the peanut genetic breeding method. The method adopts radiation mutation to create a new specie. The method comprises the following steps: the embryo of a mature seed is used as the radiation material; after the mutation, the mutated embryo is used to perform tissue culture and obtain a tissue culture regeneration seedling, and the tissue culture regeneration seedling is grafted in a field by using a seedling as rootstock. The beneficial mutation rate of the new specie obtained by the method is up to more than 30%. By using the method, the problem of the traditional method that the beneficial new peanut specie is difficult to create is solved, and the method has remarkable social and economic benefits.

The invention provides a genetic algorithm by employing guided local search for a multi-objective optimization problem. The algorithm is used for the field of flexible job-shop scheduling. A flexible job-shop scheduling problem belongs to an NP-Hard problem, optimization of multiple objectives often needs to be faced in real production, and the objectives are in interaction and conflict. The genetic algorithm aims at solving the problems of too fast convergence, insufficient population diversity and over-high calculation cost of enumerating all neighborhood solutions caused by continuous cross breeding of close relatives in a genetic operation in the genetic algorithm in the prior art. According to the algorithm, a procedure of calculating a crossover rate and a mutation rate before genetic crossover and mutation and a procedure of searching a movable process and a feasible position by using the guided local search are designed for these problems; and through introduction of the two procedures, the calculation cost is reduced while algorithm premature is avoided. The algorithm is high in practicability and can be well used for actual job-shop scheduling.

The invention discloses a kit for synchronously detecting 12 mutation hotspots of the Chinese population phenylketonuria PAH (phenylalanine hydroxylase) gene. In the kit, 12 loca on the Chinese population PAH gene are used as detection objects; an amplification primer and an extension primer are respectively designed for the mutation type of each locus; each destination section is subjected to PCR amplification and mark extension at the same time; and the gene types of the 12 mutation hotspots are obtained at the same time through capillary electrophoresis analysis. The invention provides a phenylketonuria PAH gene mutation screening kit which is simple, has the advantages of high flux, high performance, low cost and high detection accuracy, is suitable for the Chinese people and suitable for the group screening of the Chinese population phenylketonuria.