142 results about "Mutation screening" patented technology

The present invention provides oligonucleotide constructs, sets of such oligonucleotide constructs, and methods of using such oligonucleotide constructs to provide validated sequences or sets of validated sequences corresponding to desired ROIs. Such validated ROIs and constructs containing these have a wide variety of uses, including in synthetic biology, quantitative nucleic acid analysis, polymorphism and/or mutation screening, and the like.

The present invention is directed to methods and compositions for use in screening nucleic acid populations for nucleic acid polymorphisms. The methods, referred to generally as ValiGeneSM Mutation Screening, Peptide-Linked (VGMS-PL) methods, are specifically designed for high-throughput genotype mapping and gene expression analysis of animal and plant nucleic acids without requiring a PCR amplification step. In particular, the methods of the invention utilize oligonucleotide probes labeled with distinguishable and identifiable peptide tags, that are captured on addressable antibody arrays.

The invention relates to a method for preparing high purity galacto-oligosaccharides, comprising the following product separation and purification steps: Aspergillus oryzae fermentation, ceramic membrane ultrafiltration, nanofiltration separation and the like. In the method, the Aspergillus oryzae separated from the soil is adopted as an original strain and the Aspergillus oryzae BLB-21 (with preservation number of CGMCC No.2951), a high efficiency transformed strain obtained through mutation screening in the laboratory, is directly used to ferment high concentration lactose solution, thus avoiding the steps of enzyme purification and the like in the process of preparing the galacto-oligosaccharides by an enzyme method and saving time and labor. The high purity galacto-oligosaccharides are obtained by ultrafiltration and nanofiltration separation, the process is ideal, the conditions are mild and the method has extensive industrial production prospect.

A (Rhizopus Oryzae)ME-F11, its mutation screening method and method for fermenting fumaric acid are disclosed. The preserving code is CCTCC NO: M 207066. The fumaric acid concentration reaches to 90-120g/L, total yield reaches to 90-95 wt% . It's simple and can be used for industrial production.

The invention discloses a combined primer for screening human deafness gene mutation. The combined primer comprises a combined forward primer and a combined reverse primer, wherein the combined forward primer consists of two parts namely a joint part P1 sequence and a forward primer sequence, and the combined reverse primer sequentially consists of a joint part P2 sequence, a label sequence and a reverse primer sequence. The invention also discloses a kit for screening the human deafness gene mutation. The invention also discloses applications of the combined primer and the kit in preparation of a reagent for screening the human deafness multi-gene mutation. A primer containing the label (Barcode) sequence is used for differentiating different samples, and at least 32 samples can be detected by performing a method provided by the invention once, so that combined primer can be widely applied to general investigation of deafness susceptible genes. The price of the combined primer disclosed by the invention is far lower than that of imported instruments or reagents, so that the sequencing cost is greatly reduced and then the detection cost is also greatly reduced.

The invention discloses a human K-ras (K-rat sarcoma) gene mutation parting type fluorescent quantitation PCR (polymerase chain reaction) detecting reagent kit and a detecting method. The detecting reagent kit comprises PCR mixed reaction liquid, a peptide nucleic acid probe, a taqman-MGB (minor groove binder) probe, an inner control forward and reverse primer and probe, an external control forward and reverse primer and probe and a positive reference article, wherein the inner control forward and reverse primer and probe is used for K-ras gene mutation detection, and the external control forward and reverse primer and probe is used for the K-ras gene mutation detection. The PNA (peptide nucleic acid) technology and the taqman-MGB technology are combined, relevant ARMS (amplification refractory mutation system) primers are designed, PNA can be stably combined with wild K-ras genes 12 or 13 codon sequences, only mutation specimens can be amplified, and the K-ras gene mutation can be detected. The method has the advantages that the speed is high, simplicity and convenience are realized, the specificity is good, the sensitivity is high, and the method can be used for the clinical K-ras gene mutation screening.

The invention relates to primers and probes for detecting human K-ras gene mutation as well as use method thereof, relating to detection of gene mutation. In the invention, the fluorescent quantitative PCR technology and the fluorescence dual ring probes technology are combined to develop a multiple real-time fluorescent PCR method capable of fast detecting gene mutation and design the relevant primers. The method is used for detecting mutation of the twelfth and the thirteenth codons of the K-ras gene, mutant type primers designed specific to different mutation types are respectively adopted to detect samples to be detected, only the mutation type samples can be smoothly amplified to obtain double-strand DNA products which can be combined with DNA probes to send out fluorescence signals to be detected. The invention can detect mutate type genes with content being 1/1000 in the samples to be detected, has the advantages of high speed, convenience and simplicity, safety, high sensitivity, high throughput and low cost and can be applied to K-ras gene mutation screening of numerous clinical samples.

The invention discloses organophosphorus degrading enzyme mutants and coding genes and application thereof. Site-directed mutation screening is performed on original enzyme coding genes to obtain three organophosphorus degrading enzyme mutants according to the spatial structure characteristic of organophosphorus degrading enzyme OPHC2 by computer-aid molecular design, and the amino acid sequences of the three mutants are shown by SEQ ID NO.2, SEQ ID NO.4 and SEQ ID NO.6 respectively. Compared with the original enzyme, the organophosphorus degrading enzyme mutants improve the ethyl-parathion degradation specific activities by 2.45 times, 2.56 times and 4.51 times, and the degradation capabilities (kcat/Km value) of the organophosphorus degrading enzyme mutants are 2.7 times, 3.3 times and 4.2 times that of the original enzyme. Enzymatic property analysis shows that the enzymatic property of the organophosphorus degrading enzyme mutants is not changed compared with the original enzyme, and the both have good heat resistance.

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 relates to a strain for producing L-lysine, which is classified and named as Escherichiacoli NT1003 delta Met delta Thr and is collected in China Center for Type Culture Collection (CCTCC) on May 30, 2013, and the collection number is CCTCC NO: M2013239. The invention further provides a mutation screening method of the strain and a method for producing the L-lysine by utilizing the strain. After continuous passage is performed on the Escherichiacoli NT1003 delta Met delta Thr strain for 7 times, the content of the L-lysine in a fermentation solution is about 60g/L and has no significant change, so that the genetic stability is good.

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.

The invention discloses a universal strategy for efficiently improving enzyme thermodynamics stability. The strategy is specifically implemented in the way of taking a catalytic residue of zymophore as the center, selecting a high-flexibility residue 10 angstroms away from the center at most as a target locus according to the B-factor value in an enzyme crystal structure, and obtaining a mutant with enzyme thermodynamics stability improved through mutation screening. Under the guidance of the strategy, the thermal stability of candida rugosa lipase 1 with a highly complicated structure is improved remarkably. By means of stability change results of protein (LipA) with the smallest structure and moderately complicated protein (CalB), it is found that the success rate of enzyme mutation can be increased remarkably through mutation of residues with high B-factor 6-10 angstroms away from the catalytic residue. Meanwhile, through system analysis of a mutant structure-function relationship, a foundation is laid for deep research of enzyme region stabilization law and development of enzyme stabilization technique.

The invention relates to a strain for generating phospholipase D with high and stable yield by utilizing physical and chemical mutation, belonging to the technical field of microbial mutation. The strain is characterized in that the phospholipase D is prepared by fermenting streptomyces chromofuscus of high yield phospholipase D, which is obtained by taking the streptomyces chromofuscus as an original strain and sequentially carrying out composite mutation of ultraviolet rays and lithium chloride and composite mutation of atmosphere cold plasmas and the lithium chloride on a monospore bacterial suspension thereof. The invention has the advantages that the streptomyces chromofuscus of the high yield phospholipase D is obtained through mutation screening, thereby providing a high yield strain with higher application value to industrial production of the phospholipase D, laying a good foundation for fermentation culture and application of the phospholipase D, and also providing a good and feasible method for microbial mutation breeding. The invention adopts a mutation and screening method, has the advantages of simple operation, high efficiency and the like, and has a practical value on screening the strain with an industrial application value.

The invention discloses a selenium-rich yeast, a preparing method and an application. The preparing method includes the following steps that Candida ethanolica is subjected to nitrogen ion-implantation mutagenesis, and is subjected to mutation screening for multiple times, and an Candida ethanolica mutant strain tolerating high-concentration sodium selenite is obtained; a seed culturing solution is prepared, the seed culturing solution is inoculated in a fermentation medium and fermented to the logarithmic phase, the sodium selenite is additionally added into a fermented product, the mixture continues to be fermented and cultured, and a fermented product is collected; fermented liquid is subjected to centrifugal washing and dried, and then a faint-yellow powdery product is obtained. The selenium-rich yeast has a large amount of amino acid and small peptide, and is extremely and easily digested and absorbed by animals, organic selenium in the yeast can be fully absorbed by living bodies along with absorption of the amino acid and the small peptide, and the digestion using rate of the organic selenium is greatly increased. Under the condition of the equivalent additive amount, the selenium-rich yeast has the advantage of being obvious in production performance aspect compared with similar products, or under the condition of the equivalent production performance, the additive amount can be reduced, and the cost is reduced.

The invention belongs to the medicine and biotechnology field and relates to a mutation screening probe of SLC25A13 gene and application thereof. The invention provides the mutation screening probe of the SLC25A13 gene and the nucleotide coding sequence of the probe is shown in SEQ ID NO 4. The invention also provides application of the probe, namely a mutation screening method of the SLC25A13 gene. The method comprises the following steps: using the genome DNA in the sample as the template and adding the forward primer and reverse primer of the mutation site of the SLC25A13 gene and the marked normal sequence probe and mutation screening probe to carry out polymerase chain reaction; and determining whether the sample is mutant by detecting the marker in the reaction product. The probe has the characteristics of simple and direct operation, time saving, intuitive and definite result judgment, lower cost and pollution reduction.

The invention relates to POU3F4 mutant gene which has 499C> T mutation, which is relative to human genetic deafness. The invention provides a reagent kit for detecting POU3F4 gene 499 C>T mutation, and a detection process which is used to diagnose the reason and kind of deafness generation through detecting whether patients have the mutation genes. The mutant gene and the detection process are beneficial for the POU3F4 mutation screening work of deafness patients which is carried out on clinics, and provides services for the diagnosis and treatment of deaf patients.

The invention discloses bacillus subtilis producing high-temperature-resistance alpha-amylase. The bacillus subtilis Li-2013-02 is preserved in the China General Microbiological Culture Collection Center (CGMCC) on July 15, 2013, and has a preservation number of CGMCC No.7926. The strain is prepared from bacillus subtilis Li-2010 producing high-temperature-resistance alpha-amylase by means of ultraviolet ray-lithium chloride-diethyl sulfate combined mutation screening, has the characteristics of strong acid resistance, heat resistance and high enzyme production activity, and has a wide application prospect.

The invention discloses a white rot Inonotus hispidus mutant strain T906 for laccase production and a preparation method thereof. The mutant strain T906 (CoriolopsisgallicaT906) mentioned in the invention is preserved in China Center for Type Culture Collection. The preservation number is CCTCC NO: M2011317, the preservation date is September 15, 2011 and it is preserved in Wuhan University, Wuhan, China. Wild Inonotus hispidus (preservation number being cfcc88387) purchased from China Forestry Culture Collection Center is used as an original strain, and the mutant strain is obtained by ultraviolet and nitrosoguanidine combined mutation screening. Under an optimum condition, the laccase production of the mutant strain reaches 213U/mL. The mutant strain has mycelial morphology and cultural characteristics which are obviously different from the original strain, does not degenerate after serial passages for over 30 times and has genetic stability.

The present invention discloses a congenital adrenal hyperplasia gene screening kit. The congenital adrenal hyperplasia gene screening kit comprises a PCR amplification primer mixed liquor used for detecting 17 mutation sites including CYP 21A2 gene; an extended primer mixed liquor for the above 17 mutation sites; dNTP; 10*RCR buffer; 25 mM Mg2+ ions; Fast StartTaq enzyme; purification reagent composed of SAP enzyme, Exon I enzyme and a matched buffer; SNaPshot Multiplex mixed liquor; single-site homozygous and heterozygous mutation positive and negative control DNA; and deionized water. According to the technical scheme of the invention, a large number of researches and practices show that, the mutation sites of the congenital adrenal hyperplasia gene in the Chinese population are screened and combined. By utilizing the kit, 17 mutation sites of the specific congenital adrenal hyperplasia gene are simultaneously subjected to multiplex nested PCR amplification, labeling and extending, so that the defects of the existing mutation screening method are overcome. The kit is simple in operation and low in cost. The detection throughput and the accuracy of the detection result are greatly improved compared with the prior art.

The invention belongs to the technical field of biological engineering and synthetic biology, and particularly relates to a multi-fragment deoxyribose nucleic acid (DNA) series connection recombination assembly method based on BT1 integrase and mutation recognition sites thereof. In the method, sixteen pairs of mutational integrase recognition sequences are provided through mutation screening and verification, on the basis, a complete set of related carriers is constructed and comprises a framework carrier and seven TA cloning vectors, the framework carrier can realize escherichia coli-streptomyces coelicolor shuttle, the seven TA cloning vectors comprise mutation substrates and are used for DNA element construction and screening, and a concrete method for multi-fragment external series connection recombination reaction is provided. The method of the invention is simple, fast and efficient and can be used for the multi-fragment fast series connection recombination assembly, and the amplification of series connection products is realized through escherichia coli transformation. The method can be widely used for combination and assembly of standard modules and elements in the synthetic biology.

This disclosure teaches high throughput mutation screening methods allowing simultaneous analysis of multiple genetic regions of interest and sensitive detection of very low frequency mutation(s) by the use of a universalized approach. Methods comprise treating RNA:DNA heteroduplexes of interest with a ribonuclease, sequence extension by an RNA-primed DNA polymerase, ligation with a blocking adapter, and differential sequence fill-in followed by single-strand-specific nuclease digestion to permit full-length sequence extension and subsequent ligation with a tagged reporter adapter solely in mutants filled in with a complementary deoxyribonucleotide triphosphate. By forming tagged mutant-dual adapter hybrids or mutant-triple adapter hybrids, the detection and/or quantification of mutants may be directed to the commonly shared tag(s) or flanking adapter sequences for signal detection/enhancement or sequence amplification in all different mutants regardless of the source or the number of mutations involved, thereby avoiding the tremendous effort of multiple target-specific sequence amplifications. Methods may be performed wholly or partially in solution, on solid phase media, in large scale, adapted for automated or semi-automated analysis, and any combinations thereof.

The invention discloses lactococcus lactis ZF625 which is preserved in Guangdong Province Microbiological Culture Collection Center on 12 December 2018 with the preservation number GDMCC No:60522. Theinvention further discloses the use of lactococcus lactis ZF625 to food fermentation. Through mutation screening, the lactococcus lactis ZF625 which is resistant to ethanol and high in yield of lactic acid is obtained; when the lactococcus lactis ZF625 is used for brewing vinegar by a liquid state fermentation method, the lactic acid content in the vinegar can be significantly increased, the flavor of the vinegar can be improved, the irritant of the sourness of the vinegar can be reduced, and the mellow sense can be promoted.

The invention discloses a detection method for mutation screening of disease-causing genes of Parkinson's disease (PD). The detection method comprises the following steps: (1) adding dd H2O into a synthesized MIPs probe to prepare a probe solution with the concentration of 100mu M; (2) sucking 1mu l of the probe solution in step (1) by utilizing a pipettor; then putting the probe solution into a 1.5ml new EP pipe; sufficiently and uniformly mixing; marking a pipe wall as mixed-MIPs by utilizing a Marker pen; (3) preparing an MIPs probe phosphorylation activation system and carrying out activation on the mixed-MIPs; (4) diluting a mixed-MIPs probe; (5) preparing an MIPs target capturing reaction system; (6) preparing an MIPs enzyme digestion reaction system; (7) carrying out PCR (PolymeraseChain Reaction) amplification reaction; (8) carrying out polyacrylamide gel electrophoresis and magnetic bead purification; (9) carrying out high-throughput sequencing on a mixed library; (10) carrying out Sanger sequencing verification. By adopting the detection method for the mutation screening of the disease-causing genes of the Parkinson's disease, the mutation screening of the 18 PD disease-causing genes of the Parkinson's disease can be finished, and the detection method has important significance on researches of super-early diagnosis and PD pathogenesis of potential PD patients.

The invention discloses a detection method for folic acid metabolism-related gene mutation loci. The detection method comprises the steps that a sample DNA is extracted; specific primers used for amplifying folic acid metabolism-related gene segments are designed; the single-tube multi-PCR amplification reaction is carried out so as to obtain target fragments; single-base extension primers used for the folic acid metabolism-related gene segments are designed; single base extension is carried out on the folic acid metabolism-related gene fragmentx; and nucleic acid mass spectrometry analysis iscarried out so as to measure the target DNA sequences. The detection method has the advantages that folic acid metabolism-related gene mutation screening can be effectively carried out on patients with colorectal cancer, the individual can be helped to fully understand the hereditary condition of the individual, and a clinical and scientific research foundation is laid for carrying out personalized accurate diagnosis and treatment on the patients. The invention further relates to a detection kit for the folic acid metabolism-related gene mutation loci.