86results about How to "Improve sequencing efficiency" patented technology

The invention discloses a DNA (Deoxyribose Nucleic Acid) sequencer which comprises a supporting table, a plurality of vibration dampers, a vibration damping plate, a reaction bin assembly, a CCD (Charge Coupled Device) camera, a two-dimensional regulation supporting device and a medicament supply assembly, wherein the vibration damping plate is connected with the supporting table by a plurality of vibration dampers; the reaction bin assembly is fixedly arranged on the vibration damping plate and is used for performing the DNA sequencing reaction; the CCD camera is used for acquiring an optical signal; the two-dimensional regulation supporting device is used for supporting the CCD camera; and the medicament supply assembly is arranged on the supporting table and is used for providing reagents and buffer solution for the reaction bin assembly. According to the DNA sequencer disclosed by the invention, by the arrangement of a plurality of reaction bins and the matching of the two-dimensional regulation supporting device capable of carrying out two-dimensional regulation and the medicament supply assembly capable of supplying the reagents for a plurality of reaction bins, the aim of simultaneously performing a plurality of reactions is fulfilled, a plurality of samples can be simultaneously sequenced and the DNA sequencing efficiency is greatly improved.

The invention discloses a control system for a DNA (Deoxyribose Nucleic Acid) sequencer, which comprises a PLC (Programmable Logic Controller), controllers of first and second servo motors, first and second peristaltic pumps and driving motors of first and second multipass reversing valves, wherein the controllers of the first and second servo motors are electrically connected with the PLC; the first and second peristaltic pumps are respectively and electrically connected with the PLC and are used for receiving control signals of the PLC; a CCD (Charge Coupled Device) camera is electrically connected with the PLC and is used for receiving a control signal of the PLC; and a plurality of sensors are respectively and electrically connected with the PLC and are used for sending position signals of the CCD camera to the PLC. According to the control system disclosed by the invention, a reagent supply assembly of the DNA sequencer with a plurality of reaction bins timely and accurately supplies reagents and buffer solution for a plurality of reaction bins and the CCD camera can be ensured to timely read an optical signal in each reaction bin, and therefore, the aim of simultaneously performing a plurality of reactions is fulfilled, so that a plurality of samples can be simultaneously sequenced and the DNA sequencing efficiency is greatly improved.

The invention relates to a multiplex-PCR three-round amplification method, a primer and a kit thereof and application of the method in establishment of next-generation sequencing platform libraries. In the first two rounds of a PCR, a specific primer with the low concentration is consumed as much as possible, so that the differences of different amplicons are reduced and the homogeneity of multiplex-PCR products is improved; when the third round of the PCR is performed, adapter primers carrying different bar code sequences are added to mark different templates, the different adapter primers carry same universal amplification primers, therefore, it is guaranteed that the amplification efficiency of the different templates is consistent, and the differences of different template amplification products are reduced. Accordingly, not only is the problem that the homogeneity of ordinary multiplex-PCR amplification is poor solved, but also establishment of the sequencing libraries can be quickly and conveniently completed.

The invention discloses a fluid control device of gene sequencing. The fluid control device of gene sequencing comprises a reagent assembly, a first multiported valve, a first three-port valve, a gene sequencing chip and a drive assembly; the reagent assembly is connected with the gene sequencing chip through the first multiported valve and the three-port valve; due to the fact that a first gene sequencing passage and a second gene sequencing passage are arranged in the gene sequencing chip, reagents in the reagent assembly can automatically flow into the first gene sequencing passage and the second gene sequencing passage to conduct reaction and fluorescence image acquisition, when fluorescent sequencing reaction is conducted in the first gene sequencing passage, fluorescence image acquisition can be conducted in the second gene sequencing passage, and the gene sequencing time can be effectively reduced, that is to say, the gene sequencing efficiency is effectively increased. For the fluid control device of gene sequencing, the gene sequencing cost is reduced, and the gene sequencing efficiency is increased.

The invention provides a set of DNA index and application thereof to construction and sequencing of mate-paired indexed library, and the DNA index has a sequence selected from SEQ ID NO:1-24. The invention also provides a method for construction and sequencing of the mate-paired index library. The method employs two independent sequencing reactions to realize mixed sequencing on a plurality of mate-paired indexed libraries in a singe sequencing chip, so as to accelerate high flux sequencing, and reduce time, reagent cost and output cost of unit data.

The invention relates to the field of genetic engineering, in particular to a gene sequencing device and a gene sequencing system. The gene sequencing device comprises a sample preparation component, a liquid conduction component, a sample loading component, a temperature control component, an image acquisition component and a control component. The gene sequencing system comprises a sample preparation control module, a liquid conduction control module, a reaction control module, a temperature control module, an image acquisition control module and a control module. By adopting the gene sequencing device and the gene sequencing system, the integration from sample preparation to data processing can be realized, and full automation of gene sequencing can also be realized. Meanwhile, the high-pass and low-cost gene sequencing can be realized.

The invention discloses a method for simultaneously sequencing a plurality of nucleic acid samples, which comprises: respectively performing adaptor ligation, first PCR amplification, terminal phosphorylation, and DNA ligation in order of the plurality of nucleic acid samples; mixing the plurality of DNA ligation products; adding a base A at the 3'-terminal of the mixed DNA ligation products; connecting the ligation products with the 3'-terminal added with the base A to a sequencing interface; performing second PCR amplification of the sequencing interface-connected products to obtain a mixed sequencing library of the plurality of nucleic acid samples; sequencing the mixed sequencing library; and classifying the sequencing results to respectively obtain respective nucleic acid sequences of the plurality of nucleic acid samples. The method of the invention can simultaneously perform sequencing of a plurality of nucleic acid samples, is accurate in sequencing results, good in repeatability, low in cost, and high in efficiency, makes full use of sequencing flux, and is especially suitable for blood free DNA samples.

The invention provides a construction method for a simplified genome next generation sequencing library based on double enzyme digestion and a kit. Aiming at defects of an existing construction method for the double enzyme digestion simplified genome next generation sequencing library, the double enzyme digestion combined range is expanded, and excessive dependence on expensive instruments of constructing the simplified genome library is reduced, the library construction flow path is simplified, library construction cost is reduced, the sequencing efficiency is improved, and meanwhile the technology is easy and flexible to operate and easier for researchers to master and can be realized in a common molecule lab. The construction method is particularly suitable for miniature or medium-scale labs needing to conduct SNP molecular marker development, genetic map construction, population genetics research, phylogeny biological research and the like on a great number of species with incomplete reference genomes. The construction method has good practical application value and application prospects in the fields of molecular breeding of agriculture, conservation biology and evolutionary biology.

The invention relates to a sequencing data analysis system. The analysis system comprises a file renaming module, a quality control module, a sequence comparison module, a mutation detection module, amutation annotation module, a scoring and grading module, a filtering module and a mutation comment remarking module. According to the system, mutations can be detected and annotated from fastq-formatted data of a sequencing original lower computer according to a single sample or family samples, and the mutations can be scored; and after quality control, a file containing all the mutations of thesamples and annotation information and scoring information of the mutations as well as a file containing all rare mutations of the samples and annotation information and scoring information of the rare mutations are obtained finally, so that it is convenient to mine the information in sequencing data more quickly, accurately and comprehensively.

The invention relates to a gene sequencer, a liquid path system and an automatic detection method thereof. The liquid path system of the gene sequencer comprises a power mechanism, a tray, an extraction mechanism, a reaction chip and a waste liquid bottle. The power mechanism comprises an injection pump and a selecting valve connected to the injection pump; the extraction mechanism comprises a rotary valve and a plurality of extraction needles, and the rotary valve is connected to the selecting valve to form a reaction channel. The plurality of extraction needles are arranged to be inserted into reagent slots in the tray, separately. The reaction channel is connected to any one extraction needle through the rotary valve. The reaction chip is arranged on the reaction channel, and the wasteliquid bottle is connected to the selecting valve to form a waste liquid channel. The liquid path system of the gene sequencer is simple in structure. By switching the rotary valve, the injection pumpcan suck different reagents to the reaction chip for a biochemical reaction. Compared with a conventional three-dimensional mechanical arm, the rotary valve is controlled to switch more simply, so that the sequencing efficiency can be improved.

The invention provides DNA single-molecule sequencing system and apparatus based on multicolor-fluorescence reversible termination nucleotide. The DNA single-molecule sequencing system comprises a primer, a DNA template to be tested and multicolor-fluorescence reversible termination nucleotide reagents. The primer is fixed on a surface of a flow cell reactor; after hybridization of the sequencingprimer with the DNA template to be tested, the primer is extended by using the multicolor-fluorescence reversible termination nucleotide reagents; and thus, sequence information of the DNA template tobe tested can be obtained by detecting fluorescence signals of the extended primer. Localizing fluorescent marker is not required for the 3'-terminal of the DNA template to be tested. By extending fluorescence of a reactant as localizing fluorescence of the next extension in sequencing cycle or adopting a localizing fluorescent marker fixed on the surface of the flow cell reactor as localizing fluorescence, the DNA single-molecule sequencing system requires no localizing fluorescent marker at the 3'-terminal of the DNA template to be tested; so that, the problem of location information loss caused by quenching is effectively avoided. Thus, sequencing reading length can be further and greatly extended with error rate reduced.

The invention relates to circRNA and application thereof in preparation of a cervical cancer diagnostic reagent, in particular to application of hsa_circ_0000069 in preparation of a cervical cancer diagnostic reagent. Further mining and integrated analysis are carried out on sequencing data of cervical cancer, a biological information regulation network of the interaction of circRNA-miRNA-mRNA iscreated by means of a biological information method, and the key molecules hsa_circ_0000069 and hsa-miR-125b-5p which are involved in regulation of cervical cancer are searched for. A new molecular diagnostic marker for diagnosis of cervical cancer is provided, and a research basis for gene detection of clinical diseases is offered.

The invention relates to a cervical cancer molecular marker and an application thereof, and particularly relates to an application of hsa_circ_0020594 and a related gene hsa-let-7c-5p thereof in diagnosing cervical cancer. The inventor further excavates, integrates and analyzes the sequencing data of the cervical cancer, finds out the key molecule hsa_circ_0020594 participating in the regulation and control of the cervical cancer and the related gene has-let-7c-5p, draws a biological information regulation network, thereby providing a new molecular diagnostic marker for the diagnosis of the cervical squamous cell carcinoma.

The invention provides an azo linkage unit based fluorescence labeled nucleotide and applications thereof. The structural formula of the nucleotide is shown in a formula VI in the specification, wherein fluorescein is selected from one of BODIPY, rhodamine, coumarin, xanthene, cyanin, pyrene, phthalocyanine, Alexa, Squaring dye, a combination producing energy transfer dyes and derivatives thereof; R1, R2, R3 R4 and R6 are various substituent groups, R5 is a substituent group except -C2H5, and R1, R2, R3, R4, R5 and R6 are not H simultaneously; and n is an integer between 0 and 10. Compared with the prior art, a kind of new azo linkage unit based reversible terminals is synthesized according to the invention; the kind of reversible terminals can realize high-efficiency shearing under moderate conditions, so that the kind of reversible terminals can be used for DNA sequencing; and raw materials required by synthesis are simple and easy to obtain, and reactions in the synthetic process are all conventional chemical reactions, therefore, the nucleotide can be subjected to large-scale promotion and application.

The invention discloses a key microbial functional genome detection method in a pepper peeling process. According to the key microbial functional genome detection method in the pepper peeling process, from the perspective of micro-ecosystem, pepper epidermis microorganisms in the peeling process are taken as an object; metagenome DNA of pepper epidermis bacteria is extracted and purified and a variable region sequence is amplified, and then the variable region sequence of the epidermis bacteria is read with the application of a high-throughput sequencing technology; by virtue of a bioinformatics platform, the sequence is subjected to quality control and species annotation and function prediction are completed, and on the basis, a microorganism species, which plays a vital role in the pepper peeling process, in fresh fruit epidermis and a core metabolic pathway thereof are comprehensively and objectively revealed depending on a statistical method. The detection method provided by the invention simplifies experimental operations and improves a sequencing efficiency, so that the development of metagenomics is greatly promoted.

The invention belongs to the technical field of genotyping detection method, and in particular, relates to a multi-PCR-SBT genotyping method for an ABO antigen of a human erythrocyte blood type system. The invention also relates to a multi-PCR-SBT reagent for ABO antigen genotyping of the human erythrocyte blood type system. The reagent and method provided by the invention can be used as an independent and widely used identification method, solve the problem of rapid and accurate typing of an ABO antigen system, and exert the characteristics of high-throughput operation and accurate results ofmulti-PCR-SBT on ABO genotyping. Relevant applications in the fields of clinical blood transfusion medicine research and genetics can be highly valued, and important practical significance are achieved in research departments, and pharmaceutical research and reagent development departments.

The invention relates to a medium-throughput gene expression analysis method based on a second-generation test platform. The medium-throughput gene expression analysis method comprises steps as follows: corresponding PCR (polymerase chain reaction) primers are divided into two groups according to the relative expression quantity of multiple genes, and the concentration proportion of primers is adjusted in the groups; competitive templates of standard substances and a competitive template of a to-be-tested sample are established, and three rounds of multiple competitive PCRs (polymerase chain reactions) are performed; reaction products of the three rounds of reactions are mixed to serve as a second-generation testing platform library for computer sequencing, data are extracted and analyzed, and the difference of relative expression quantities of multiple genes in different samples can be obtained. On the basis of the high-speed-developed second-generation sequencing platform, a target template and an internal reference template are accurately and rapidly quantified, expression difference analysis of five target genes can be performed on multiple samples simultaneously, the procedure is simple and easy to implement, and the cost is low; when multiple gene expression differences of multiple samples are required to be analyzed, the method can be a high-accuracy, moderate-throughput and low-cost method.

The invention relates to a hypoxic-ischemic brain damage diagnosis target and application. Hypoxic-ischemic encephalopathy of the newborn is one of the common reasons of child nervous system impairment, and no effective diagnosis methods for the hypoxic-ischemic encephalopathy exist currently. The invention performs circRNA transcriptome sequencing on a hypoxic-ischemic brain damage model, discovers 9 circRNA closely related to the hypoxic-ischemic brain damage through bioinformatics analysis and verifies the 9 circRNA. The invention provides a potential biological marker for the diagnosis ofthe hypoxic-ischemic brain damage and lays a foundation for the clinical gene detection of the hypoxic-ischemic brain damage.

The invention relates to circRNA_14707 and an application thereof in molecular assistant breeding. By means of high-throughput sequencing on expression conditions of circRNA in subcutaneous fat tissues of a big white pig and a Laiwu pig and integrated analysis performed by combining previous miRNA and mRNA of a laboratory, a result shows that differential expressions of the circRNA_14707 and related miRNA are quite obvious and the circRNA_14707 and related miRNA can serve as molecular markers for detecting the subcutaneous fat contents of the pigs. The circRNA_14707 has a very good applicationprospect in the fields of predicating or assisted predicting pork quality and breeding pigs with different muscle qualities and the like.

The invention discloses ctDNA library construction and sequencing data analysis methods for simultaneously detecting common mutations of various liver cancers. The library construction method and thesequencing data analysis method have the following advantages: firstly, various mutation forms of liver cancers are simultaneously detected under the condition that capture is not needed; secondly, the methods are suitable for efficient capturing in an ultra-small target area; thirdly, the library can support 10 to 20 times of detection; fourthly, the DNA barcode is connected to an initial ctDNA molecule during the library construction process and the flow is analyzed by cooperating with a biological information analysis process to realize high-specificity detection of ctDNA low-frequency mutation; and fifthly, the library can be used for PCR hot spot detection and capture method sequencing at the same time, the false positive mutation can be filtered effectively by using the added DNA barcode to realize duplex-based high-specificity sequencing. The methods have the important clinical significance for early screening, disease tracking, curative effect evaluation, prognosis prediction and the like of liver cancers.

The invention relates to a double-shielding box-type DNA sequencer with a portal. The double-shielding box-type DNA sequencer comprises a reaction record subsystem, a reagent supply system, a control system and a sequencer box bearing the systems, the reaction record subsystem comprises a reaction bin component, a movement platform component, a movement control unit, a CCD camera and a platform supporting component, a reaction chip component is arranged in the reaction bin component, and reaction liquid enters a reaction chip for chemical reaction to generate visible light under action of the control system. The double-shielding box-type DNA sequencer comprises the portal and a turnable reaction bin double-shielding structure, the reaction chip can be loaded under double protection, and the portal itself has extremely high stability; after long-time use, the portal does not deform, and the CCD camera can be stabilized at a same point for testing.

The invention relates to circRNA_26852 and application thereof and in particular relates to circRNA_26852 and application thereof in pig molecule auxiliary breeding. Expression of circRNA in subcutaneous fat tissue of large white pigs and Laiwu pigs is identified and analyzed for a first time, hoping to find potential circRNA related to adipogenic differentiation and lipid metabolism. Results showthat with the combination of circRNA_26852 and miRNA such as ssc-miR-486 and ssc-miR-874, expression of related genes in related signal paths of fat deposition and lipid metabolism can be regulated and controlled. With the combination of circRNA_26852 and miRNA, expression of related genes in signal paths is regulated and controlled, a potential regulation and control function can be brought intoplay in fat deposition and lipid metabolism, and a base can be made for development of improved varieties of livestock.

The invention relates to novel circRNA related to intramuscular fat and application thereof, in particular to circRNA_08840 related to intramuscular fat and new application of an interaction gene ssc-miR-339-3p of circRNA_08840. The two kinds of typical pigs which are large white pigs and Laiwu pigs are selected as research objects, a biological information science method is used for conducting sequencing and data analyzing on circRNA, miRNA and an interaction relation of circRNA and miRNA in the intramuscular fat of the two kinds of pigs, several quite good intramuscular fat content marks arefound, and experimental results show that circRNA_08840 and the interaction gene ssc-miR-339-3p of circRNA_08840 are remarkable in expression difference of intramuscular fat tissue of the two kinds of typical pigs, and novel circRNA is expected to breed pigs with different meat qualities as a mark in molecule auxiliary breeding.

The invention discloses a fluorescence-labeled azo-modified nucleotide and application thereof in DNA sequencing. The nucleotide has a structural formula as shown in a formula I, wherein R1, R2, R3, R4, R5 and R6 may not synchronously be H, and m and n are integers of 0 to 10. Compared with the prior art, novel reversible terminators based on azo connection units are synthesized; a denaturing PAGE gel, i.e. a sequencing gel proves that the reversible terminators can only extend a reversible terminator at a time and has extension efficiency of 100% when a template is a plurality of continuous same basic groups; thus, under the action of a reducing agent, highly-efficient, rapid and full shearing can be realized, and great potential and value applied in the DNA sequencing are obtained; meanwhile, raw materials needed by synthesis of the azo-modified nucleotide are simple and easily available, and synthetic process is a conventional chemical reaction and can be used for popularization and application.

The invention provides a gene sequencing light-path system. The gene sequencing light-path system disclosed by the embodiment of the invention comprises two excitation light sources, the two excitation light sources are running simultaneously, two light beams are combined by a first beam combiner, and then the combined light beam passing through a main beam splitter mirror, after being focused by a focusing microscope, is irradiated on a to-be-detected sample. Therefore, the gene sequencing light-path system can achieve an effect of simultaneously exciting visible light with various specific wavelengths, thereby improving the sequencing efficiency of the gene sequencing light-path system.

The invention relates to a DNA sequencer, comprising a reaction recording subsystem, a reagent supply system and a control system, and a sequencer box which bears the above systems, wherein the control system is disposed at the rear of the reaction recording subsystem disposed a left area of the box and controls the reaction subsystem and the reagent supply system to act according to a preset time sequence; the control system controls the reagent supply system to transfer a reagent to the reaction recording subsystem, the reagent flows through a reaction chip disposed in a reaction chamber module to react, a CCD (charge coupled device) camera of the reaction recording subsystem collects photos in a reaction process and acquires a measured DNA sequence by analyzing. During a DNA sequencing process, the sequencer necessarily features continuation and controllability during reagent supply and sequencing reaction processes, sequencing reaction and reagent supply equipment are provided with a stable, closed operating environment; the whole structure is stable, and sequencing efficiency is high.

The invention relates to the field of automatic control, and provides a sequencing control system. The sequencing control system comprises a first sequencing response unit, a first figure collection unit, a second sequencing response unit, a second figure collection unit and a control unit, wherein the first sequencing response unit is used for control over the sequencing response of a first sequencing response chamber; the first figure collection unit is used for control over signal detection and collection for the first sequencing response chamber by a figure collection component to obtain signal data; the second sequencing response unit is used for control over sequencing response of a second sequencing response chamber; the second figure collection unit is used for control over signal detection and collection for the second sequencing response chamber by the figure collection component to obtain signal data; and the control unit is used for control over multiple times of cycle operation of the first sequencing response unit and the first figure collection unit, and also use for control over multiple times of cycle operation of the second sequencing response unit and the second figure collection unit. The sequencing control system can enable rapid sequencing to be achieved, and capable of improving the use ratio of the sequencing control system.

The invention relates to a circRNA and application thereof in detecting intramuscular fat. A circRNA-seq technology and a bioinformatics method are utilized for identifying and analyzing circRNA in the intramuscular fat of Laiwu pigs and large white pigs, circRNA_23437 relevant to regulation and control of pig fat decomposition is found out, the mechanism of the circRNA in regulating fat decomposition and the fat metabolism function is further known, and a theoretical basis is laid for improving the quality of pork.

The invention relates to a box type DNA sequencing instrument with a frame type shock absorption structure. The box type DNA sequencing instrument comprises a response record subsystem, a reagent supply system, a control system and a sequencing instrument box body for loading the systems, wherein the sequencing instrument box body comprises a main supporting component for supporting a bottom part, a side part, a back part and a division part between a left box body area and a right box body area of the box body; the main supporting component is formed by splicing steel plates, is fixed by splicing or bolt fastening, and comprises a base component, a top plate component, a supporting frame component which is connected with the base component to form a side frame of the box body, and a buffer system mounting seat; and the supporting frame component comprises a left side vertical plate component and a middle vertical plate component for separating a left box body and a right box body. The box type DNA sequencing instrument with the frame type shock absorption structure has a frame structure composed of the steel plate and has a strong shock absorption effect; sequencing reaction and reagent supply equipment should have a stable and closed operating environment; and the box type DNA sequencing instrument is stable in overall structure and high in sequencing efficiency.