80 results about "Restriction fragment" patented technology

The invention provides methods for determining genome-wide sequence variations associated with a phenotype of a species in a hypothesis-free manner. In the methods of the invention, a set of restriction fragments for each of a sub-population of individuals having the phenotype are generated by digesting nucleic acids from the individual using one or more different restriction enzymes. A set of restriction sequence tags for the individual is then determined from the set of restriction fragments. The restriction sequence tags for the sub-population of organisms are compared and grouped into one or more groups, each of which comprising restriction sequence tags that comprise homologous sequences. The obtained one or more groups of restriction sequence tags identify the sequence variations associated with the phenotype. The methods of the invention can be used for, e.g., analysis of large numbers of sequence variants in many patient samples to identify subtle genetic risk factors.

The invention provides a method for constructing a high resolution physical map of a polynucleotide. In accordance with the invention, nucleotide sequences are determined at the ends of restriction fragments produced by a plurality of digestions with a plurality of combinations of restriction endonucleases so that a pair of nucleotide sequences is obtained for each restriction fragment. A physical map of the polynucleotide is constructed by ordering the pairs of sequences by matching the identical sequences among the pairs.

The present invention relates to methods and apparatuses for characterization of single polymers. In particular, the invention relates to methods and apparatuses for determination of the velocities of single elongated polymers. Center-of-mass velocity, center-to-center velocity, end-to-end velocity and rise-time velocity are determined using time-correlated measurements of single elongated polymers in two or more detection zones. The invention also relates to methods of determinating lengths and molecular masses of single polymers and to methods of determining the distance between landmarks on a single polymers based on their velocities. The invention further relates to methods of single-molecule DNA restriction fragment analysis.

The present invention relates to a high throughput method for the identification and detection of molecular markers wherein restriction fragments are generated and suitable adaptors comprising (sample-specific) identifiers are ligated. The adapter-ligated restriction fragments may be selectively amplified with adaptor compatible primers carrying selective nucleotides at their 3′ end. The amplified adapter-ligated restriction fragments are, at least partly, sequenced using high throughput sequencing methods and the sequence parts of the restriction fragments together with the sample-specific identifiers serve as molecular markers.

The present invention relates to methods and apparatuses for characterization of single polymers. In particular, the invention relates to methods and apparatuses for determination of the velocities of single elongated polymers. Center-of-mass velocity, center-to-center velocity, end-to-end velocity and rise-time velocity are determined using time-correlated measurements of single elongated polymers in two or more detection zones. The invention also relates to methods of determinating lengths and molecular masses of single polymers and to methods of determining the distance between landmarks on a single polymers based on their velocities. The invention further relates to methods of single-molecule DNA restriction fragment analysis.

The present invention relates to a high throughput method for the identification and detection of molecular markers wherein restriction fragments are generated and suitable adaptors comprising (sample-specific) identifiers are ligated. The adapter-ligated restriction fragments may be selectively amplified with adaptor compatible primers carrying selective nucleotides at their 3′ end. The amplified adapter-ligated restriction fragments are, at least partly, sequenced using high throughput sequencing methods and the sequence parts of the restriction fragments together with the sample-specific identifiers serve as molecular marker.

The present invention provides methods for rapid forensic analysis of mitochondrial DNA by amplification of a segment of mitochondrial DNA containing restriction sites, digesting the mitochondrial DNA segments with restriction enzymes, determining the molecular masses of the restriction fragments and comparing the molecular masses with the molecular masses of theoretical restriction digests of known mitochondrial DNA sequences stored in a database.

The present invention provides methods for rapid forensic analysis of mitochondrial DNA by amplification of a segment of mitochondrial DNA containing restriction sites, digesting the mitochondrial DNA segments with restriction enzymes, determining the molecular masses of the restriction fragments and comparing the molecular masses with the molecular masses of theoretical restriction digests of known mitochondrial DNA sequences stored in a database.

The invention provides methods and compositions for amplifying selected polynucleotides, especially selected subsets of restriction fragments. Generally, methods of the invention are implemented by ligating adaptors containing at least one promoter sequence to such fragments under conditions that promote the formation of closed single stranded or double stranded structures, which are capable of serving as cyclical templates for transcription.

Disclosed is a method of detecting bacteria in a biological sample, especially a blood sample, without the need for extensive sub-culturing of the sample. Nucleic acid present within the sample is isolated and bacterial DNA specifically amplified using primers that uniquely prime the amplification of 16s rRNA-encoding nucleic acid. The amplicons are then digested with an endonuclease to yield a restriction fragment length profile for the biological sample. The restriction fragment length profile for the biological sample is then compare to a database of profiles made using cultures of known bacterial species. A match between the sample profile and the database quickly identifies the bacteria present in the sample.

A method for determining a genome sequence comprising the steps of digesting the genome with at least one first restriction endonuclease, ligating at least one adaptor to the restriction fragments of the first subset, selectively amplifying the first set of adaptor-ligated restriction fragments using a first primer combination wherein at least a first primer contains a first selected sequence at the 3′ end of the primer sequence, comprising 1-10 selective nucleotides, repeating these steps with at least a second primer combinations wherein the primer contains a different second selected sequence, fragmenting each of the subsets of amplified adaptor-ligated restriction fragments to generate sequencing libraries, determine the nucleotide sequence of the fragments, aligning the sequence of the fragments in each of the libraries to generate contigs, repeating these steps for one second and / or further restriction endonucleases, aligning the contigs obtained for each of the second and / or further restriction endonucleases to provide for a sequence of the genome.

Disclosed is a method of detecting bacteria in a biological sample, especially a blood sample, without the need for extensive sub-culturing of the sample. Nucleic acid present within the sample is isolated and bacterial DNA specifically amplified using primers that uniquely prime the amplification of 16s rRNA-encoding nucleic acid. The amplicons are then digested with an endonuclease to yield a restriction fragment length profile for the biological sample. The restriction fragment length profile for the biological sample is then compare to a database of profiles made using cultures of known bacterial species. A match between the sample profile and the database identifies the bacteria present in the sample.

The invention relates to a molecule marking method for pig backfat thickness property. The method comprises: extraction of pig genome DNA, design of a long-chain acyl-coenzyme Asynthetase 1 (ACSL1) gene seventh exon primer, amplification in vitro and genotype detection. For the first time, it is discovered that the polymorphism of the long-chain acyl-coenzyme Asynthetase 1 (ACSL1) gene seventeenth exon has significant correlation with the pig backfat thickness, and a method for detecting the restriction fragment polymorphism at mutation sites of the seventeenth exon is established. The molecule marking method is applicable to auxiliary selection on backfat thickness property during breeding of pigs, helps to realize early-stage breeding selection of boars, and even helps to accurately select breeding just when pigs are born, so that the generation interval is shortened, and the selection progress of the backfat thickness property is accelerated; and the method is simple in operation, condition requirements during a polymerase chain reaction are low, the length of amplified fragments is relatively short (292 bp), amplification is relatively easy, and the method helps to improve amplification efficiency and accuracy of genotype determining.

Methods for detecting genomic rearrangements are provided. In one embodiment, methods are provided for the use of paired end tags from restriction fragments to detect genomic rearrangements. Sequences from the ends of the fragments are brought together to form ditags and the ditags are detected. Combinations of ditags are detected by an on-chip sequencing strategy that is described herein, using inosine for de novo sequencing of short segments of DNA. In another aspect, translocations are identified by using target specific capture and analysis of the captured products on a tiling array.

The invention provides methods for determining genome-wide sequence variations associated with a phenotype of a species in a hypothesis-free manner. In the methods of the invention, a set of restriction fragments for each of a sub-population of individuals having the phenotype are generated by digesting nucleic acids from the individual using one or more different restriction enzymes. A set of restriction sequence tags for the individual is then determined from the set of restriction fragments. The restriction sequence tags for the sub-population of organisms are compared and grouped into one or more groups, each of which comprising restriction sequence tags that comprise homologous sequences. The obtained one or more groups of restriction sequence tags identify the sequence variations associated with the phenotype. The methods of the invention can be used for, e.g., analysis of large numbers of sequence variants in many patient samples to identify subtle genetic risk factors.

The invention belongs to the technical field of genetic engineering, and provides a method for acquiring chicken whole genome high-density SNP marker sites. The method comprises the following steps: (1) predicting distribution of restriction fragments acquired from double-restriction chicken genomes of EcoRI and MseI; (2) designing a universal joint, a barcode joint and a PCR amplification primer according to the distribution characteristics of the restriction fragments of EcoRI and MseI; (3) constructing a simplified genome sequencing library; (4) sequencing by virtue of the library constructed in the step (3) through a computer; and (5) according to a sequencing result, acquiring SNP marker sites. The method disclosed by the invention provides a universal strategy for the construction of a whole genome high-density SNP map by virtue of double-restriction GBS for different varieties of chickens, so that the cost on acquiring every SNP marker site is reduced by an order of magnitude compared with a conventional chip technology; and the method is stable in technology and high in repeatability.

The present invention relates to a molecular marking method for pig litter size. The method comprises the steps of pig genome DNA extraction, exon 6 primer design of sex hormone binding globulin (SHBG) genes, amplification in vitro and genotype detection. The present invention first discovers pig SHBG genes, discovers the obvious relationship between the polymorphism of exon 6 and pig litter size, and develops a restriction fragment enzyme digestion polymorphism detection method for detecting exon 6 mutational sites. The method can be used for the auxiliary selection of litter size during pig breeding, can realize the early selection of parent pigs and even can accurately select parent pigs when the pigs are born. Generation interval is largely shortened, and selection progress is accelerated. The method has the advantages of simple operation, low condition requirements in polymerase chain reaction processes, short amplification fragment length (217bp), easy amplification, high amplification efficiency and accurate genotype judgement.

The present invention relates, in general, to the genetics of viruses. In particular, the present invention provides methods of distinguishing between serotypes of avian infectious bronchitis virus based on restriction fragment length polymorphisms derived from the region of the S1 gene of IBV, peptides derived from the restriction fragments, related primers for polymerase chain reaction, and certain restriction length polymorphism patterns.

The invention discloses a staphylococcus aureus multiple PCR-MIX rapid detection kit and a detection method thereof; the detection kit comprises PCR MIX; wherein, the PCR MIX contains 2*PCR buffer, 1.0-3.0mmol / L of MgCl2, 180-220mu mol / L of dNTP each, heat-resisting deoxyribonuclease nuc gene primer of which the base sequence is SEQ NO.1 and the concentration of SEQ NO.1 is 100-600nmol / L, plasma-coagulase clfA gene primer of which the base sequence is SEQ NO.3 and the concentration of SEQ NO.4 is 100-600nmol / L, SmaI restriction fragment specific sequence primer of which the base sequence is SEQ NO.5 and the concentration of SEQ NO.6 is 100-600nmol / L, 20-60U of Taq enzyme and bromophenyl blue dye. The multiple PCR-MIX rapid detection kit and the detection method provided by the invention have simple configuration, the detection kit and the detection method can be used in industrialized production; the detection method is sensitive and has short detection cycle and strong maneuverability, so that the method can be widely applied in the fields of food hygiene, environmental monitoring, etc.

Methods of analyzing DNA to identify regions of the genome that are methylated in a genomic sample are disclosed. In one aspect genomic DNA is fragmented using a restriction enzyme with a degenerate recognition site, methylated restriction fragments are separated from unmethylated fragments by affinity purification. The complexity of the methylated fragments is reduced by amplification of a subset of the fragments using adaptors that ligate to a subset of the fragments. The amplified product is fragmented, labeled and hybridized to an array of probes. The hybridization pattern is analyzed to determine methylation status of cytosines.

A method for determining a genome sequence comprising the steps of digesting the genome with at least one first restriction endonuclease, ligating at least one adaptor to the restriction fragments of the first subset, selectively amplifying the first set of adaptor-ligated restriction fragments using a first primer combination wherein at least a first primer contains a first selected sequence at the 3' end of the primer sequence, comprising 1-10 selective nucleotides, repeating these steps with at least a second primer combinations wherein the primer contains a different second selected sequence, fragmenting each of the subsets of amplified adaptor-ligated restriction fragments to generate sequencing libraries, determine the nucleotide sequence of the fragments, aligning the sequence of the fragments in each of the libraries to generate contigs, repeating these steps for one second and / or further restriction endonucleases, aligning the contigs obtained for each of the second and / or further restriction endonucleases to provide for a sequence of the genome.

Method for de novo whole genome sequencing based on a (sequence-based) physical map of a DNA sample clone bank based on end-sequencing tagged adapter-ligated restriction fragments, in combination with sequencing adapter-ligated restriction fragments of the DNA sample wherein the recognition sequence of the restriction enzyme used in the generation of the physical map is identical to at least part of the recognition sequence of the restriction enzyme used in the generation of the DNA sample.

The invention provides methods and compositions for amplifying selected polynucleotides, especially selected subsets of restriction fragments. Generally, methods of the invention are implemented by ligating adaptors containing at least one promoter sequence to such fragments under conditions that promote the formation of closed single stranded or double stranded structures, which are capable of serving as cyclical templates for transcription.

The invention relates to a process for the controlled amplification of at least one part of a starting DNA containing a plurality of restriction sites for a determined specific restriction endonuclease, and of which at least part or its nucleic acid is unknown.Application of this process to human, animal or plant DNA fingerprinting, to identification of restriction fragment length polymorphisms.Kit for the application of the process.

The present invention provides a method for identifying a plurality of pairs of interacting proteins and plasmids for use in the method. The pair of plasmids is adapted for use in a modified two hybrid system wherein wherein each plasmid comprises a recombinase recognition site. The method comprises the steps of providing cDNAs encoding test polypeptides, inserting the cDNAs into the first and second plasmids, recombining the first and second plasmids to obtain recombined plasmids, isolating and digesting the recombined plasmids, ligating the restriction fragments to a universal adapter to provide a pool of digested fragments flanked by a universal adapter, selecting and amplifying desired sequences, forming concatamers from the amplified sequences, and sequencing the concatamers to determine the nucleotide sequences encoding a plurality of pairs of interacting proteins.

A method for de novo whole genome sequencing based on a (sequence-based) physical map of a DNA sample clone bank based on end-sequencing tagged adapter-ligated restriction fragments, in combination with sequencing adapter-ligated restriction fragments of the DNA sample wherein the recognition sequence of the restriction enzyme used in the generation of the physical map is identical to at least part of the recognition sequence of the restriction enzyme used in the generation of the DNA sample.

Early detection of pathogens during asymptotic stage can be useful for timely need based adoption of available methodologies to prevent establishment of the disease and associated yield loss. Beside this, it would be useful for predicting crop productivity and monitoring presence of pathogens in the samples being traded internationally by quarantine departments. Our innovation of designing universal primer to PCR amplify fungal rDNA sequences, restriction digestion of PCR products with HinFl enzyme and analysis of the restriction fragments through PathDec tool leads identification and detection of almost all the fungal pathogens of apple even before visual appearance of the disease symptoms in less than 6 hours.

The present invention relates to a high throughput method for the identification and detection of molecular markers wherein restriction fragments are generated and suitable adaptors comprising (sample-specific) identifiers are ligated. The adapter- ligated restriction fragments may be selectively amplified with adaptor compatible primers carrying selective nucleotides at their 3' end. The amplified adapter-ligated restriction fragments are, at least partly, sequenced using high throughput sequencing methods and the sequence parts of the restriction fragments together with the sample-specific identifiers serve as molecular markers.

The invention discloses a method for identifying the traditional Chinese medical material corn cervi pantotrichum based on PCR-RFLP. A reagent kit used for identifying the traditional Chinese medical material corn cervi pantotrichum comprises a primer pair shown in (a) or (b) and restriction enzyme DdeI or isoschizomers of the restriction enzyme DdeI, wherein (a) is the primer pair composed of two lines of single-stranded DNA molecules shown in a sequence 1 and a sequence 2 in a sequence table, (b) is the primer pair which is composed of two lines of single-stranded DNA molecules shown in the sequence obtained by replacing and / or deleting and / or adding one or several nucleotides shown in the sequence 1 and the sequence 2 in the sequence table and is identical with the primer pair in (a) in function. According to the method, cytochrome b gene sequences of the traditional Chinese medical material corn cervi pantotrichum and counterfeit drug of the traditional Chinese medical material corn cervi pantotrichum are analyzed, different SNP loci of the traditional Chinese medical material corn cervi pantotrichum and the counterfeit drug of the traditional Chinese medical material corn cervi pantotrichum are obtained, identification primers and the restriction enzyme are designed, and the traditional Chinese medical material corn cervi pantotrichum and the counterfeit drug of the traditional Chinese medical material corn cervi pantotrichum are accurately identified through the restriction fragment length polymorohism polymerase chain reaction technology.

The present invention provides methods and kits useful for enriching, identifying and quantifying methylated DNA3 particularly hypermethylated CpG islands by digesting a sample with a methylation-sensitive restriction endonuclease and capturing methylated restriction fragments with a methyl-binding capture reagent. The methods of the invention may be used in the detection of cancer, particularly detection of prostate cancer.