CN110904209A - DNA library for detecting and diagnosing nystagmus disease-causing gene and application thereof - Google Patents
DNA library for detecting and diagnosing nystagmus disease-causing gene and application thereof Download PDFInfo
- Publication number
- CN110904209A CN110904209A CN201911113267.0A CN201911113267A CN110904209A CN 110904209 A CN110904209 A CN 110904209A CN 201911113267 A CN201911113267 A CN 201911113267A CN 110904209 A CN110904209 A CN 110904209A
- Authority
- CN
- China
- Prior art keywords
- nystagmus
- library
- disease
- genes
- dna
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
- C12Q1/6874—Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation
-
- C—CHEMISTRY; METALLURGY
- C40—COMBINATORIAL TECHNOLOGY
- C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
- C40B40/00—Libraries per se, e.g. arrays, mixtures
- C40B40/04—Libraries containing only organic compounds
- C40B40/06—Libraries containing nucleotides or polynucleotides, or derivatives thereof
- C40B40/08—Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/118—Prognosis of disease development
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Molecular Biology (AREA)
- Genetics & Genomics (AREA)
- Biochemistry (AREA)
- Analytical Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Biotechnology (AREA)
- Immunology (AREA)
- Microbiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Pathology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention relates to a DNA library for detecting and diagnosing nystagmus disease-causing genes by a targeted high-throughput sequencing technology and application thereof, wherein the library comprises 674 nystagmus disease-causing genes. According to the invention, 674 nystagmus disease-causing genes are preferably selected, a probe pool is designed, a target region library aiming at the 674 nystagmus disease-causing genes is established, the library is sequenced by using a high-throughput sequencing technology, disease-causing mutation is searched, and genetic and molecular biological bases are provided for clinical diagnosis. The 674 genes related to the invention comprise almost all pathogenic genes of genetic diseases taking nystagmus as clinical expression, such as X-linked congenital nystagmus, eye skin albinism, ataxia, congenital amaurosis, cataract and the like, and have important significance and clinical value for diagnosis, differential diagnosis and accurate treatment of the nystagmus.
Description
Technical Field
The invention belongs to the technical field of biological medicines, and particularly relates to a DNA library for detecting and diagnosing nystagmus pathogenic genes by a targeted high-throughput sequencing technology and application thereof.
Background
Nystagmus (NY) is an involuntary, rhythmic, back-and-forth oscillating movement of the eye. The directions are classified into horizontal type, vertical type, rotation type, etc., and the horizontal type is common, and the nystagmus direction is usually expressed by the fast phase direction, and the fast phase is compensatory to recover the movement of the fixation position, called nystagmus for short. Nystagmus can be found in the early postnatal period, and the incidence rate is about 1/1000-1/1500. In the case of eyeball tremor, it seriously impairs visual function, and both eyes vibrate continuously or the appearance is affected with compensation of head position and head swing, which causes great trouble to the life of patients. Meanwhile, nystagmus can be caused by diseases of a visual system, extraocular muscles, labyrinths of inner ears and a central nervous system, sometimes nystagmus is not an independent disease but a clinical manifestation of some diseases, even a unique clinical manifestation in early stage of diseases, such as albinism, optic neuropathy and the like, so once nystagmus manifestation is found, the cause of the disease should be determined in time, and targeted treatment and consultation are adopted.
Nystagmus has a simple phenotype, but the hidden causes of the nystagmus are very complex, and if only symptomatic treatment is performed, the treatment effect is poor and the treatment time is delayed. Recent clinical research results show that the hereditary disease caused by many pathogenic gene variations is the very murder of nystagmus, so that the nystagmus can be diagnosed only by using gene detection. At present, the traditional Sanger sequencing method is mostly adopted for detecting gene mutation in a clinical laboratory, and if a plurality of pathogenic genes of nystagmus are simultaneously detected, the workload is huge, the detection efficiency is low, more importantly, precious DNA samples are wasted, the detection cost of gene diagnosis is obviously increased, and the large-scale application of the gene diagnosis in clinical molecular diagnosis is severely restricted. Therefore, there is a need to find a new method for detecting nystagmus disease-causing gene mutation, which can improve the diagnosis accuracy, reduce the cost and labor intensity, and improve the timeliness.
Disclosure of Invention
The invention aims to provide a DNA library for detecting and diagnosing nystagmus disease-causing genes by a targeted high-throughput sequencing technology, which can improve the diagnosis accuracy and reduce the cost and the labor intensity.
The second object of the present invention is to provide the use of said DNA library.
The purpose of the invention is realized by the following technical scheme: a DNA library for diagnosing nystagmus based on a high-throughput sequencing technology, the library comprising 674 nystagmus virulence genes, wherein the 674 nystagmus virulence genes are as follows:
A2ML1、AARS、AARS2、ABCA4、ABCB7、ABHD12、ABHD5、ACO2、ACOX1、ADAMTSL4、ADAR、ADD3、ADGRG1、ADGRV1、ADSL、AFG3L2、AGBL5、AGK、AHI1、AIPL1、ALDH18A1、ALG13、ALG2、ALMS1、ALX4、ANK1、ANKRD11、ANO10、ANOS1、ANTXR1、AP3B1、AP3D1、AP4E1、APC、APTX、ARHGEF18、ARID1A、ARID1B、ARID2、ARL13B、ARL2BP、ARL6、ARMC9、ARNT2、ARX、ASAH1、ASPA、ATAD3A、ATCAY、ATF6、ATM、ATN1、ATP13A2、ATP1A2、ATP1A3、ATP6V1E1、ATXN1、ATXN10、ATXN2、ATXN3、ATXN7、ATXN8OS、B3GLCT、B9D1、BAZ1B、BBIP1、BBS1、BBS10、BBS12、BBS2、BBS4、BBS5、BBS7、BBS9、BCOR、BCS1L、BDNF、BEAN1、BEST1、BLOC1S6、BMP4、BRAF、BRCA2、BRIP1、BUB1B、CABP4、CACNA1A、CACNA1F、CACNA1G、CACNA2D4、CACNB4、CAPN1、CASK、CAV1、CC2D2A、CCDC141、CDH3、CDHR1、CEP104、CEP120、CEP164、CEP290、CEP41、CEP78、CERKL、CHD7、CHMP1A、CHN1、CISD2、CLCN7、CLDN16、CLDN19、CLIP2、CLN5、CLP1、CLRN1、CNGA1、CNGA3、CNGB1、CNGB3、CNNM4、COG4、COL11A1、COL18A1、COL25A1、COL2A1、COQ2、COQ8A、COX10、COX15、CPLX1、CRB1、CRX、CRYAA、CRYBA4、CRYBB1、CRYBB2、CRYGC、CRYGD、CSPP1、CTBP1、CTDP1、CYP1B1、CYP7B1、DAB1、DARS、DARS2、DCX、DGUOK、DHCR24、DHCR7、DHDDS、DKK1、DLAT、DNAJC12、DNM1L、DPAGT1、DPM1、DPP6、DPYD、DTNBP1、DUSP6、EBP、ECHS1、EDNRB、EEF2、EFNB1、EGR2、EIF2S3、ELN、ELOVL4、ELOVL5、ELP4、EPG5、ERCC1、ERCC2、ERCC3、ERCC4、ERCC5、ERCC6、ERCC8、ERMARD、ESCO2、EXOSC3、EXT2、EYS、FA2H、FAM161A、FANCA、FANCB、FANCC、FANCD2、FANCE、FANCF、FANCG、FANCI、FANCL、FANCM、FBN1、FBXL4、FEZF1、FGF14、FGF17、FGF8、FGFR1、FGFRL1、FLRT3、FMR1、FOXC1、FOXE3、FOXL2、FOXP1、FOXRED1、FRMD7、FRRS1L、FSCN2、FXN、GABRD、GALC、GAN、GBA、GBA2、GDF3、GDF6、GFM1、GJA1、GJA8、GJB1、GJC2、GLRX5、GMPPA、GMPPB、GNAS、GNAT1、GNAT2、GNB1、GNB3、GNB5、GPR143、GPR179、GRID2、GRK1、GRM1、GRM6、GTF2E2、GTF2IRD1、GUCA1B、GUCY2D、HARS、HDAC8、HECW2、HESX1、HGSNAT、HIBCH、HIKESHI、HMGB3、HMX1、HPS1、HPS3、HS6ST1、HSD17B10、HSD17B4、HSPD1、HTRA1、HYLS1、IARS2、IDH3B、IFT122、IFT140、IFT172、IFT27、IFT43、IFT52、IGBP1、IL17RD、IMPDH1、IMPG2、INPP5E、INPP5K、IQCB1、ITPR1、KAT6B、KCNAB2、KCNC3、KCND3、KCNJ13、KCNV2、KDM6A、KIAA0556、KIAA0586、KIDINS220、KIF1A、KIF1C、KIF2A、KIF5A、KIF7、KISS1R、KIZ、KLHL7、KMT2A、KMT2D、KRAS、L1CAM、L2HGDH、LAMA1、LAMB2、LARGE1、LCA5、LETM1、LIG4、LIM2、LIMK1、LMNB1、LONP1、LRAT、LRIT3、LRP4、LRP5、LYRM7、LYST、LZTFL1、LZTR1、MAB21L2、MAD2L2、MAFB、MAG、MAK、MAN2B1、MAP2K1、MAP2K2、MARS2、MBTPS2、MC1R、MCOLN1、MECR、MERTK、MITF、MKKS、MKS1、MMACHC、MMADHC、MME、MOCS1、MOCS2、MPC1、MPDU1、MPLKIP、MPV17、MPZ、MRE11、MTFMT、MTPAP、MTR、MTRR、MVK、MYO5A、MYO7A、NAA10、NADK2、NAGA、NALCN、NANS、NARS2、NAXE、NDP、NDUFA10、NDUFA11、NDUFA12、NDUFA2、NDUFA9、NDUFAF1、NDUFAF2、NDUFAF3、NDUFAF4、NDUFAF5、NDUFAF6、NDUFB11、NDUFB3、NDUFB9、NDUFS1、NDUFS2、NDUFS3、NDUFS4、NDUFS6、NDUFS7、NDUFS8、NDUFV1、NDUFV2、NEK2、NELFA、NEU1、NFIX、NHS、NIPBL、NMNAT1、NPHP1、NPHP4、NR2E3、NR2F1、NRAS、NRL、NSD1、NSD2、NSMF、NSUN2、NT5C2、NUBPL、NUP62、NYX、OCA2、OCLN、OCRL、OFD1、OPA1、OPA3、OPHN1、OPN1LW、OPN1MW、OTX2、PACS1、PALB2、PAX2、PAX6、PCYT1A、PDE6A、PDE6B、PDE6C、PDE6G、PDE6H、PDGFRB、PDP1、PDZD7、PEX10、PEX11B、PEX12、PEX13、PEX14、PEX16、PEX19、PEX2、PEX26、PEX3、PEX5、PEX6、PEX7、PHF21A、PHF6、PHGDH、PHYH、PIGN、PIGT、PIK3R5、PITX2、PLA2G6、PLEKHG2、PLK4、PLP1、PMM2、PMP22、PMPCA、PNPLA6、POC1B、POLG、POLR3A、POLR3B、POMGNT1、PORCN、PRCD、PRDM16、PRKCG、PRKD1、PRNP、PROK2、PROKR2、PROM1、PRPF31、PRPF4、PRPF6、PRPF8、PRPH2、PRPS1、PRRT2、PRSS12、PRX、PTPN11、PTPN22、PURA、PYCR2、RAB18、RAD21、RAD51、RAD51C、RAF1、RAP1A、RAP1B、RARS、RASA2、RBP3、RD3、RDH12、RECQL4、REEP6、RERE、RFC2、RGR、RHO、RIT1、RLBP1、RNASEH2C、RNASET2、RNF216、ROBO3、RP2、RP9、RPE65、RPGRIP1、RPGRIP1L、RRAS、RTN4IP1、RUBCN、SACS、SAG、SALL4、SAMD9L、SCN1A、SCN8A、SDCCAG8、SDHA、SDHAF1、SDHD、SEMA3A、SEMA3E、SEMA4A、SEPSECS、SERPINI1、SETD5、SETX、SH3TC2、SIL1、SIM1、SIN3A、SIX6、SKI、SLC12A6、SLC16A2、SLC17A5、SLC18A3、SLC19A1、SLC19A2、SLC19A3、SLC1A3、SLC24A1、SLC24A5、SLC25A20、SLC25A4、SLC29A3、SLC2A1、SLC33A1、SLC35A2、SLC38A8、SLC39A8、SLC45A2、SLC4A11、SLC52A2、SLC6A19、SLC7A14、SLC9A1、SLC9A6、SLX4、SMARCA4、SMARCB1、SMARCE1、SMC1A、SMC3、SNIP1、SNRNP200、SNX10、SNX14、SOS1、SOS2、SOST、SOX10、SOX11、SOX2、SOX3、SPAST、SPATA5、SPATA7、SPG11、SPG7、SPRY4、SPTBN2、SQSTM1、SRD5A3、SRY、STUB1、STX16、SURF1、SYNE1、SYT14、TACR3、TAF1、TAF2、TBL2、TBP、TCIRG1、TCTN1、TCTN2、TENM3、TFAP2A、TIMMDC1、TINF2、TMEM126A、TMEM126B、TMEM138、TMEM216、TMEM231、TMEM237、TMEM240、TMEM67、TNFRSF11A、TNFSF11、TOE1、TOPORS、TRAF3IP1、TREX1、TRIM32、TRIM44、TRPM1、TTBK2、TTC19、TTC8、TTPA、TTR、TUBB3、TUBGCP4、TUBGCP6、TUFM、TULP1、TWNK、TYR、TYRP1、UBA5、UBE2T、UBE3A、UBE3B、UCHL1、UNC80、UROC1、USH2A、VHL、VLDLR、VPS13A、VPS13B、VWA3B、WDPCP、WDR11、WDR19、WDR35、WDR73、WFS1、WHRN、WT1、WWOX、XRCC2、XRCC4、XYLT2、YAP1、YARS2、ZFYVE26、ZNF408、ZNF423、ZNF513、ZNF592。
the DNA library provided by the invention covers almost all genetic diseases taking nystagmus as clinical expression, such as X-linked congenital nystagmus, eye skin albinism, ataxia, congenital amaurosis, cataract and the like, and comprises 674 nystagmus pathogenic genes. These 674 gene selections are based on the internationally reported and recognized clinical disease gene database (OMIM database, HGMD database, ClinVar database). The 674 pathogenic gene variants can cause diseases independently or jointly to cause more serious and complex clinical phenotypes, and the invention can comprehensively sequence the 674 genes at one time for the first time.
According to 674 nystagmus disease-causing genes, a probe pool which can cover exons of the genes and is adjacent to a +/-20 bp intron region is designed, a target region library containing the 674 nystagmus disease-causing genes is established by utilizing probe target capture, the library is sequenced by utilizing a high-throughput sequencing technology, disease-causing mutation is searched, the genetic cause of nystagmus is determined, and the theoretical basis of genetics and molecular biology is provided for clinical diagnosis.
The invention also provides application of the DNA library in preparation of a diagnostic kit, wherein the kit is used for diagnosing the pathogenic gene of nystagmus.
Further, the application comprises the following steps:
1) clinical data and clinical biological samples of nystagmus patients are collected, preferably, the clinical biological samples refer to various samples from human bodies, including but not limited to peripheral blood, body fluid and tissue and organ samples from subjects, such as saliva, hair or oral mucosa of the subjects.
2) Extracting the genome DNA of the sample, preferably, the extraction method comprises a DNA extraction kit or various manual extraction methods;
3) quantifying the extracted genomic DNA and constructing a library, preferably, the quantification methods include, but are not limited to, fluorescence quantification methods and electrophoresis; wherein, the library construction comprises the following steps:
a) fragmenting genomic DNA, preferably by methods including but not limited to ultrasonication, transposase cleavage, and restriction endonuclease cleavage;
b) carrying out end repair on the fragmented genomic DNA and simultaneously carrying out 3' end addition of A;
c) ligating the product of adding A at the 3' end to a linker for amplification of the ligation-effective product, preferably from a high throughput sequencing library kit;
d) performing PCR amplification on the ligation product by using a universal primer, and adding a complete joint, wherein the universal primer is preferably from a high-throughput sequencing and library building kit;
e) using probes aiming at the 674 nystagmus disease genes to target and capture a target area, preferably, the capture method comprises but is not limited to liquid phase probe capture and solid phase chip hybridization capture;
f) washing off the uncaptured library, and only reserving the target area library;
j) obtaining a target region capture library.
4) Performing quantitative operation on the library, preferably, the quantitative method includes but is not limited to fluorescence quantitative method and electrophoresis;
5) performing high throughput sequencing of the library using a sequencing device, preferably, the sequencing device includes, but is not limited to, a Novaseq series, a Hiseq series, a Nexeseq series, a BGIseq series second generation nucleic acid sequencer;
6) and comparing the obtained sequencing data by bioinformatics, and obtaining the related information of the pathogenic site by mutation interpretation.
A diagnostic kit comprising said DNA library.
Compared with the prior art, the invention has the advantages that:
1. nystagmus is related to various genetic diseases, early symptoms are not serious and are easily ignored by patients, and when serious complications or other symptoms occur, the latest treatment opportunity is often missed, so that the determination of a pathogenic gene is the key of clinically correct diagnosis of nystagmus. The invention aims to establish a new method for quickly, accurately and high-flux detecting nystagmus disease-causing genes, thereby helping to understand pathogenesis and laying a foundation for assisting clinical diagnosis, prognosis judgment, prenatal diagnosis and accurate treatment.
2. The gene diagnosis is helpful for clinical genetic counseling, prenatal diagnosis and the like. Through clinical genetic consultation, family members can know the possibility of suffering from diseases; parents or the patients can guide the birth through gene detection, and the health of the next fetus in the future is ensured. Nystagmus is often one of the clinical phenotypes of genetic diseases and can provide accurate health and reproductive guidance only if genetic testing is performed. Finding carriers of asymptomatic young virulence genes in families also helps to develop home medical and rehabilitation programs earlier.
3. The DNA library of the application refers to a large number of documents on the basis that the inventor develops high-throughput gene detection service for many years to accumulate a large number of clinical cases of the nystagmus in China, and finally prefers 674 nystagmus pathogenic genes by adopting a disease-gene correlation screening method provided by ClinGen and other power databases, and gives consideration to comprehensiveness, accuracy and scientificity. According to the invention, 674 nystagmus disease-causing genes are preferably selected, a probe pool is designed, a target region library aiming at the 674 nystagmus disease-causing genes is established, the library is sequenced by using a high-throughput sequencing technology, disease-causing mutation is searched, and genetic and molecular biological bases are provided for clinical diagnosis. The 674 gene detection regions can detect all genetic diseases which take nystagmus as clinical manifestations, such as X-linked congenital nystagmus, eye skin albinism, ataxia, congenital amaurosis, cataract and the like, and have important significance and clinical value for diagnosis and differential diagnosis of nystagmus.
4. The invention adopts a high-throughput sequencing technology to sequence the target region capture library, and can simultaneously detect all exons and adjacent regions of 674 nystagmus pathogenic genes involved in the invention in one sequencing reaction. Compared with the traditional sequencing technology, the method has the advantages of obviously improved detection efficiency and obviously reduced cost, has great advantages in nystagmus gene excavation and pathogenic gene screening, and is an efficient, reliable and economic nystagmus pathogenic gene detection technology.
5. In conclusion, the DNA library and the application thereof have the characteristics of accuracy, flexibility, rapidness and low cost; through clinical evaluation, the invention has good auxiliary diagnostic value for the eyeball tremor.
Drawings
FIG. 1 is the quality control information for establishing a target region targeting DNA library for 674 nystagmus disease-causing genes of proband samples in example 1;
FIG. 2 is data coverage information for sequencing the target region of 674 nystagmus virulence genes of proband samples in example 1
FIG. 3 is data amount information for sequencing the target region of 674 nystagmus virulence genes of proband sample in example 1
FIG. 4 is a pedigree map of nystagmus pedigree as described in example 1, in which arrows indicate probands, solid icons indicate patients, and open icons indicate healthy individuals.
Detailed Description
The invention provides a DNA library for detecting and diagnosing nystagmus disease-causing genes and application thereof, which are further described in the following by combining specific embodiments.
Unless otherwise indicated, the techniques used in the examples are conventional and well known to those skilled in the art, and may be performed according to the fourth edition of the molecular cloning, laboratory Manual, or related information, and the reagents and products used are also commercially available. Various procedures and methods not described in detail are conventional methods well known in the art, and the sources, trade names, and components of the reagents used are indicated at the time of first appearance, and the same reagents used thereafter are the same as those indicated at the first appearance, unless otherwise specified.
Example 1:
in this embodiment, a Hiseq sequencing platform of Illumina corporation is used to detect genomic DNA of peripheral blood of a human subject, and the specific implementation steps are as follows:
1. sample source
A congenital nystagmus family from the Chinese Fujian province, wherein the predecessor is a 25-year-old male, the patient finds nystagmus from small (about 6 months old), the vision is corrected by 0.4, the refractive interstitial substance is transparent when the anterior segment of the eye is checked, the reflex of the pupil opposite side is normal, the eyeground, optic nerve and macula examination are not abnormal, the electroretinogram is not abnormal, and the history of marriage of near relatives is denied. The family has 2 men and 2 women with the same symptoms, the onset age is 6 months to 1 year, and no other obvious abnormality is found in ophthalmic examination. The family tree of this family is shown in fig. 4, with arrows pointing to probands, solid icons representing patients, and open icons representing healthy individuals. A total of 6 family members participate in the detection, including proband (sick), proband mother (sick), proband father (healthy), proband sister (healthy), proband aunt (sick) and proband epiglottis (sick), informed consent of all participants is obtained, and 10mL of venous blood (EDTA anticoagulated treatment) is collected from the elbow vein of the family member of the embodiment for detection.
2. Extraction of specimen genomic DNA:
genomic DNA extraction kit (HiPureblood) from magenta was used according to the instructions provided by the trade company&Tissue DNA Kit) genomic DNA was extracted from peripheral blood samples, the purity of the DNA was measured using Nanodrop one, and OD of the genomic DNA obtained260nm/OD280nmAll are located between 1.7 and 2.0, and the concentration of the DNA is measured by using Nanodrop one, and the concentration of the obtained genomic DNA is 50 to 100 ng/. mu.L, and the total amount is 5 to 10. mu.g.
3. Establishing a genome amplification library:
according to the instructions provided by the trade company, the Kit of KAPA company (KAPA HyperPlusLibraryPreparation Kit) is used for carrying out enzyme digestion and fragmentation, end repair, 3' end A addition, linker ligation and PCR amplification on genomic DNA, and finally, the obtained library is quantified and quality checked, and the specific implementation steps are as follows:
1) and (3) carrying out genome DNA fragmentation reaction, wherein the reaction system is as follows:
| name of reagent | Dosage (mu L) |
| Genomic DNA (100ng) | 1 |
| NF H2O | 16.5 |
| KAPA Frag Buffer | 2.5 |
| KAPA Frag Enzyme | 5 |
| |
25 |
Reaction conditions are as follows: the reaction was carried out at 37 ℃ for 12 min.
2) End repairing and 3' end adding A reaction, wherein the reaction system is as follows:
reaction conditions are as follows: reacting at 20 ℃ for 1 min; reacting at 65 ℃ for 30 min; constant temperature of 20 DEG C
3) Linker linking reaction, the reaction system is as follows:
reaction conditions are as follows: the reaction was carried out at 20 ℃ for 20 min.
4) The first PCR amplification reaction comprises the following reaction systems:
| name of reagent | Dosage (mu L) |
| DNA obtained in step 3) | 16 |
| KAPA HiFi HotStart Ready Mix(2×) | 20 |
| T5*Primer(10μM) | 1.5 |
| T8*Primer(10μM) | 1.5 |
| Total volume | 39 |
Reaction conditions are as follows: 45s at 98 ℃; (98 ℃ for 15s,60 ℃ for 30s,72 ℃ for 30 s). times.6 cycles; 1min at 72 ℃; storing at 12 deg.C.
5) DNA quantification and quality control:
measuring the concentration and the strip distribution condition of the PCR amplification product by using Nanodrop one; preparing 2% agarose gel, mixing the obtained PCR amplification product with the sample loading buffer solution, and observing the position, brightness and uniformity of the band by electrophoresis, wherein the size of the band is qualified between 200 and 800 bp.
4. Constructing a target region targeting DNA library based on probe hybridization capture:
according to the operation of the instruction provided by the merchant, the library is constructed by adopting a library construction kit of IGT company, a probe is designed according to the sequence of 674 candidate nystagmus pathogenic genes, and is synthesized and labeled by biotin, and the specific implementation steps are as follows:
1) and (3) probe hybridization:
mixing 5 mu g of PCR product obtained in the step 3 with 5 mu L of Cot-1human DNA, and carrying out vortex oscillation; adding 2.5 × Ampure XP beads according to the total volume, uniformly mixing and centrifuging, and standing for 5min at room temperature; the magnetic beads are centrifugally washed for 2 times by 80% ethanol, and are eluted by a hybridization solution for hybridization reaction, wherein the system of the hybridization solution is as follows:
hybridization conditions: reacting at 95 ℃ for 10 min; at 65 ℃ overnight
2) The probes hybridized with the sample target sequences were captured on magnetic beads by binding biotin to Streptavidin using 80. mu.L of Streptavidin-labeled magnetic beads (M-270Streptavidin beads). Washing with 200 μ L of 1 × Bead Washbuffer at 65 deg.C and room temperature for three times (3 min each), and washing the magnetic beads with 20 μ L NF H2And (4) resuspending the solution.
3) And carrying out a second PCR amplification reaction on the captured target sequence, wherein the reaction system is as follows:
| name of reagent | Dosage (mu L) |
| KAPA HiFi Hot start Readymix(2×) | 25 |
| X Gen Library Amplification primer-Ts Mix | 5 |
| DNA with magnetic beads obtained in step 2) | 20 |
| |
50 |
Reaction conditions are as follows: 45s at 98 ℃; (98 ℃ for 15s,60 ℃ for 30s,72 ℃ for 30 s). times.7 cycles; 1min at 72 ℃; storing at 4 ℃.
5) And (3) PCR product quantification and quality inspection:
measuring the concentration and the strip distribution condition of the PCR amplification product by using Nanodrop one; preparing 2% agarose gel, mixing the obtained PCR amplification product with the sample loading buffer solution, and observing the position, brightness and uniformity of the band by electrophoresis, wherein the size of the band is qualified between 200 and 800 bp.
5. Generating information analysis and variant interpretation of sequencing data:
NGS sequencing results were aligned to the human reference genome UCSC NCBI37/hg19 using Novocraft Novoalign to obtain a unique aligned sequence aligned to the genome. Variation in the 674 nystagmus disease-causing gene regions was determined using the VarScan mpileup2snp and VarScan mpileup2indel assays. Common variations in dbSNP and ExAC databases were removed using Remove Run Common Variants and Remove Global Common Variants software. The variants were then annotated using Interactive Biosoftware Alamut Batch. The database used for annotation includes: dbSNP, ExAC, 1000g, ClinVar, OMIM, etc., and utilizes the software FATHMM, FATHMMMKL, METALR, METASVM, MUTATIONASSESSOR, MUTATIONTASTERAGGGD, AGVGD, LRT, PROVEAN, SIFT to predict the variant function. According to the ACMG genetic variation classification standard and guideline, mutation sites which are meaningful for diagnosing the nystagmus are obtained through analysis.
6. Sequencing results interpretation and analysis
674 nystagmus pathogenic genes of 6 examinees are sequenced at one time by the DNA library for detecting and diagnosing the nystagmus pathogenic genes. Taking prover as an example, the size of the obtained DNA library is mainly distributed in 200-800bp through quality control analysis, the average length is 399bp, and the concentration and the fragment size meet the sequencing requirement, thereby prompting that the quality of the library construction is qualified (as shown in figure 1). The DNA library comprises 674 genes, wherein the length of a coding region is 5131171bp, the length of a non-coding region is 1189045bp, and the total length is 6320216 bp. Wherein the coding region has a 10 × coverage of 99.51%, a 20 × coverage of 99.25%, and a 50 × coverage of 94.83% (see fig. 2). A Total of 11629197 fragment read length data (Total Reads) were obtained with a match rate (Aligned) of 90.49% (see FIG. 3). The sequencing result meets the data requirement of variation interpretation. Through analysis of sequencing data, the pathogenic gene variation of nystagmus (X-linked congenital nystagmus type 1) related to the clinical phenotype of the tested person is found, and is shown in the following table:
in this example, the peripheral blood genomic DNA of the proband was detected to have a hybrid pathogenic mutation of NM _194277.2: c.443T > G (p.Leu 148) of FRMD7 gene, i.e., the 443 th base T of the coding region was replaced by G, and the nonsense mutation (p.Leu 148) resulted in a premature stop codon, which may cause protein truncation or activation of nonsense-mediated mRNA degradation, thereby affecting the function of the protein product encoded by FRMD7 gene. This truncation variation has not been reported in the literature so far, but truncation variations downstream of this variation are known to be reported as pathogenic variations (PubMed:17013395,27036142,24434814,24513357). The mutation was detected in the peripheral blood DNA of the sick proband mother, aunt and epiglottis, and was not detected in the peripheral blood DNA of the healthy proband father and sister, suggesting that the mutation was co-isolated from the clinical phenotype in the pedigree. The pathogenic variation suggests that the proband suffers from X-linked congenital nystagmus 1 type, and provides a basis for clinical diagnosis. The disease condition of the X-linked congenital nystagmus 1 type patient is mainly eye expression without other complication risks. The treatment suggestion is symptomatic treatment, such as vision protection, eye fatigue prevention, etc. For genetic counseling, since the disease is an X-linked genetic disease, the probability of 1/2 for children of female patients is that the patient is ill, children of male patients are not ill, and children are ill. It is worth mentioning that the symptoms of female patients may be very mild. (see fig. 4)
Reference throughout this specification to the description of "one embodiment," "this embodiment," "an embodiment," or the like, means that a particular feature, structure, material, or characteristic described in connection with the embodiment is included in at least one embodiment of the invention and is not specifically referred to.
It should be noted that, although the above embodiments have been described herein, the invention is not limited thereto. Therefore, based on the innovative concepts of the present invention, the technical solutions of the present invention can be directly or indirectly applied to other related technical fields by making changes and modifications to the embodiments described herein, or by using equivalent structures or equivalent processes performed in the content of the present specification and the attached drawings, which are included in the scope of the present invention.
Claims (8)
1. A DNA library for diagnosing nystagmus based on a high-throughput sequencing technology, wherein the library comprises 674 nystagmus pathogenic genes, wherein the 674 nystagmus pathogenic genes are as follows:
A2ML1、AARS、AARS2、ABCA4、ABCB7、ABHD12、ABHD5、ACO2、ACOX1、ADAMTSL4、ADAR、ADD3、ADGRG1、ADGRV1、ADSL、AFG3L2、AGBL5、AGK、AHI1、AIPL1、ALDH18A1、ALG13、ALG2、ALMS1、ALX4、ANK1、ANKRD11、ANO10、ANOS1、ANTXR1、AP3B1、AP3D1、AP4E1、APC、APTX、ARHGEF18、ARID1A、ARID1B、ARID2、ARL13B、ARL2BP、ARL6、ARMC9、ARNT2、ARX、ASAH1、ASPA、ATAD3A、ATCAY、ATF6、ATM、ATN1、ATP13A2、ATP1A2、ATP1A3、ATP6V1E1、ATXN1、ATXN10、ATXN2、ATXN3、ATXN7、ATXN8OS、B3GLCT、B9D1、BAZ1B、BBIP1、BBS1、BBS10、BBS12、BBS2、BBS4、BBS5、BBS7、BBS9、BCOR、BCS1L、BDNF、BEAN1、BEST1、BLOC1S6、BMP4、BRAF、BRCA2、BRIP1、BUB1B、CABP4、CACNA1A、CACNA1F、CACNA1G、CACNA2D4、CACNB4、CAPN1、CASK、CAV1、CC2D2A、CCDC141、CDH3、CDHR1、CEP104、CEP120、CEP164、CEP290、CEP41、CEP78、CERKL、CHD7、CHMP1A、CHN1、CISD2、CLCN7、CLDN16、CLDN19、CLIP2、CLN5、CLP1、CLRN1、CNGA1、CNGA3、CNGB1、CNGB3、CNNM4、COG4、COL11A1、COL18A1、COL25A1、COL2A1、COQ2、COQ8A、COX10、COX15、CPLX1、CRB1、CRX、CRYAA、CRYBA4、CRYBB1、CRYBB2、CRYGC、CRYGD、CSPP1、CTBP1、CTDP1、CYP1B1、CYP7B1、DAB1、DARS、DARS2、DCX、DGUOK、DHCR24、DHCR7、DHDDS、DKK1、DLAT、DNAJC12、DNM1L、DPAGT1、DPM1、DPP6、DPYD、DTNBP1、DUSP6、EBP、ECHS1、EDNRB、EEF2、EFNB1、EGR2、EIF2S3、ELN、ELOVL4、ELOVL5、ELP4、EPG5、ERCC1、ERCC2、ERCC3、ERCC4、ERCC5、ERCC6、ERCC8、ERMARD、ESCO2、EXOSC3、EXT2、EYS、FA2H、FAM161A、FANCA、FANCB、FANCC、FANCD2、FANCE、FANCF、FANCG、FANCI、FANCL、FANCM、FBN1、FBXL4、FEZF1、FGF14、FGF17、FGF8、FGFR1、FGFRL1、FLRT3、FMR1、FOXC1、FOXE3、FOXL2、FOXP1、FOXRED1、FRMD7、FRRS1L、FSCN2、FXN、GABRD、GALC、GAN、GBA、GBA2、GDF3、GDF6、GFM1、GJA1、GJA8、GJB1、GJC2、GLRX5、GMPPA、GMPPB、GNAS、GNAT1、GNAT2、GNB1、GNB3、GNB5、GPR143、GPR179、GRID2、GRK1、GRM1、GRM6、GTF2E2、GTF2IRD1、GUCA1B、GUCY2D、HARS、HDAC8、HECW2、HESX1、HGSNAT、HIBCH、HIKESHI、HMGB3、HMX1、HPS1、HPS3、HS6ST1、HSD17B10、HSD17B4、HSPD1、HTRA1、HYLS1、IARS2、IDH3B、IFT122、IFT140、IFT172、IFT27、IFT43、IFT52、IGBP1、IL17RD、IMPDH1、IMPG2、INPP5E、INPP5K、IQCB1、ITPR1、KAT6B、KCNAB2、KCNC3、KCND3、KCNJ13、KCNV2、KDM6A、KIAA0556、KIAA0586、KIDINS220、KIF1A、KIF1C、KIF2A、KIF5A、KIF7、KISS1R、KIZ、KLHL7、KMT2A、KMT2D、KRAS、L1CAM、L2HGDH、LAMA1、LAMB2、LARGE1、LCA5、LETM1、LIG4、LIM2、LIMK1、LMNB1、LONP1、LRAT、LRIT3、LRP4、LRP5、LYRM7、LYST、LZTFL1、LZTR1、MAB21L2、MAD2L2、MAFB、MAG、MAK、MAN2B1、MAP2K1、MAP2K2、MARS2、MBTPS2、MC1R、MCOLN1、MECR、MERTK、MITF、MKKS、MKS1、MMACHC、MMADHC、MME、MOCS1、MOCS2、MPC1、MPDU1、MPLKIP、MPV17、MPZ、MRE11、MTFMT、MTPAP、MTR、MTRR、MVK、MYO5A、MYO7A、NAA10、NADK2、NAGA、NALCN、NANS、NARS2、NAXE、NDP、NDUFA10、NDUFA11、NDUFA12、NDUFA2、NDUFA9、NDUFAF1、NDUFAF2、NDUFAF3、NDUFAF4、NDUFAF5、NDUFAF6、NDUFB11、NDUFB3、NDUFB9、NDUFS1、NDUFS2、NDUFS3、NDUFS4、NDUFS6、NDUFS7、NDUFS8、NDUFV1、NDUFV2、NEK2、NELFA、NEU1、NFIX、NHS、NIPBL、NMNAT1、NPHP1、NPHP4、NR2E3、NR2F1、NRAS、NRL、NSD1、NSD2、NSMF、NSUN2、NT5C2、NUBPL、NUP62、NYX、OCA2、OCLN、OCRL、OFD1、OPA1、OPA3、OPHN1、OPN1LW、OPN1MW、OTX2、PACS1、PALB2、PAX2、PAX6、PCYT1A、PDE6A、PDE6B、PDE6C、PDE6G、PDE6H、PDGFRB、PDP1、PDZD7、PEX10、PEX11B、PEX12、PEX13、PEX14、PEX16、PEX19、PEX2、PEX26、PEX3、PEX5、PEX6、PEX7、PHF21A、PHF6、PHGDH、PHYH、PIGN、PIGT、PIK3R5、PITX2、PLA2G6、PLEKHG2、PLK4、PLP1、PMM2、PMP22、PMPCA、PNPLA6、POC1B、POLG、POLR3A、POLR3B、POMGNT1、PORCN、PRCD、PRDM16、PRKCG、PRKD1、PRNP、PROK2、PROKR2、PROM1、PRPF31、PRPF4、PRPF6、PRPF8、PRPH2、PRPS1、PRRT2、PRSS12、PRX、PTPN11、PTPN22、PURA、PYCR2、RAB18、RAD21、RAD51、RAD51C、RAF1、RAP1A、RAP1B、RARS、RASA2、RBP3、RD3、RDH12、RECQL4、REEP6、RERE、RFC2、RGR、RHO、RIT1、RLBP1、RNASEH2C、RNASET2、RNF216、ROBO3、RP2、RP9、RPE65、RPGRIP1、RPGRIP1L、RRAS、RTN4IP1、RUBCN、SACS、SAG、SALL4、SAMD9L、SCN1A、SCN8A、SDCCAG8、SDHA、SDHAF1、SDHD、SEMA3A、SEMA3E、SEMA4A、SEPSECS、SERPINI1、SETD5、SETX、SH3TC2、SIL1、SIM1、SIN3A、SIX6、SKI、SLC12A6、SLC16A2、SLC17A5、SLC18A3、SLC19A1、SLC19A2、SLC19A3、SLC1A3、SLC24A1、SLC24A5、SLC25A20、SLC25A4、SLC29A3、SLC2A1、SLC33A1、SLC35A2、SLC38A8、SLC39A8、SLC45A2、SLC4A11、SLC52A2、SLC6A19、SLC7A14、SLC9A1、SLC9A6、SLX4、SMARCA4、SMARCB1、SMARCE1、SMC1A、SMC3、SNIP1、SNRNP200、SNX10、SNX14、SOS1、SOS2、SOST、SOX10、SOX11、SOX2、SOX3、SPAST、SPATA5、SPATA7、SPG11、SPG7、SPRY4、SPTBN2、SQSTM1、SRD5A3、SRY、STUB1、STX16、SURF1、SYNE1、SYT14、TACR3、TAF1、TAF2、TBL2、TBP、TCIRG1、TCTN1、TCTN2、TENM3、TFAP2A、TIMMDC1、TINF2、TMEM126A、TMEM126B、TMEM138、TMEM216、TMEM231、TMEM237、TMEM240、TMEM67、TNFRSF11A、TNFSF11、TOE1、TOPORS、TRAF3IP1、TREX1、TRIM32、TRIM44、TRPM1、TTBK2、TTC19、TTC8、TTPA、TTR、TUBB3、TUBGCP4、TUBGCP6、TUFM、TULP1、TWNK、TYR、TYRP1、UBA5、UBE2T、UBE3A、UBE3B、UCHL1、UNC80、UROC1、USH2A、VHL、VLDLR、VPS13A、VPS13B、VWA3B、WDPCP、WDR11、WDR19、WDR35、WDR73、WFS1、WHRN、WT1、WWOX、XRCC2、XRCC4、XYLT2、YAP1、YARS2、ZFYVE26、ZNF408、ZNF423、ZNF513、ZNF592。
2. use of a DNA library according to claim 1 for the preparation of a diagnostic kit, characterized in that: the kit is used for diagnosing and discovering nystagmus pathogenic genes.
3. The application according to claim 2, characterized in that it comprises the following steps:
1) extracting genomic DNA of a sample of a subject;
2) quantifying the extracted genomic DNA and constructing a library according to the following steps:
a) fragmenting the genomic DNA;
b) adding a base A to the 3' end of the fragmented genomic DNA while performing end repair;
c) connecting a product with a base A added at the 3' end with a joint;
d) performing PCR amplification on the ligation product, and adding a complete linker;
e) targeting capture of a target region using a probe directed against the 674 nystagmus virulence genes of claim 1;
f) washing off the uncaptured library, and only reserving the target area library;
j) obtaining a target region capture library;
3) performing quantitative operation on the library;
4) high-throughput sequencing;
5) and (5) analyzing the data to obtain the related information of the pathogenic site.
4. Use according to claim 3, characterized in that: the sample in the step 1) is from peripheral blood, body fluid and a tissue organ sample of a subject.
5. Use according to claim 3, characterized in that: the quantitative method in the step 2) comprises a fluorescence quantitative method and electrophoresis.
6. Use according to claim 3, characterized in that: the fragmentation method in the step a) comprises ultrasonic crushing, transposase enzyme digestion and restriction enzyme digestion.
7. Use according to claim 3, characterized in that: the quantitative method in the step 3) comprises a fluorescence quantitative method and electrophoresis.
8. A diagnostic kit comprising the DNA library of claim 1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201911113267.0A CN110904209A (en) | 2019-11-14 | 2019-11-14 | DNA library for detecting and diagnosing nystagmus disease-causing gene and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201911113267.0A CN110904209A (en) | 2019-11-14 | 2019-11-14 | DNA library for detecting and diagnosing nystagmus disease-causing gene and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN110904209A true CN110904209A (en) | 2020-03-24 |
Family
ID=69816593
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201911113267.0A Pending CN110904209A (en) | 2019-11-14 | 2019-11-14 | DNA library for detecting and diagnosing nystagmus disease-causing gene and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN110904209A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111363804A (en) * | 2020-04-29 | 2020-07-03 | 中信湘雅生殖与遗传专科医院有限公司 | Detection method, detection composition and detection kit for Joubert syndrome |
| CN111778281A (en) * | 2020-07-17 | 2020-10-16 | 四川省人民医院 | Construction method and application of retina bipolar cytopathy model |
| CN113774128A (en) * | 2021-09-24 | 2021-12-10 | 中国人民解放军东部战区总医院 | Gja8 application of gene mutation site in preparation of product for diagnosing cataract disease |
| CN117778562A (en) * | 2023-04-14 | 2024-03-29 | 湖北省妇幼保健院（湖北省妇女儿童医院） | UBE3B mutant gene, detection kit and method |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060099578A1 (en) * | 2001-08-30 | 2006-05-11 | Wallace Douglas C | Mitochondrial biology expression arrays |
| RU2005114809A (en) * | 2005-05-16 | 2006-11-20 | Саратовский государственный университет им. Н.Г. Чернышевского (RU) | METHOD FOR TREATING EYE NISTAGMA |
| CN102409095A (en) * | 2011-11-17 | 2012-04-11 | 哈尔滨医科大学 | Congenital nystagmus gene detection kit |
| CN108676865A (en) * | 2018-04-08 | 2018-10-19 | 复旦大学附属眼耳鼻喉科医院 | A kind of glaucoma of childhood related gene chip and its preparation method and application |
| US20180346981A1 (en) * | 2014-11-20 | 2018-12-06 | Massachusetts Eye And Ear Infirmary | Panel-based Genetic Diagnostic Testing for Inherited Eye Diseases |
-
2019
- 2019-11-14 CN CN201911113267.0A patent/CN110904209A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060099578A1 (en) * | 2001-08-30 | 2006-05-11 | Wallace Douglas C | Mitochondrial biology expression arrays |
| RU2005114809A (en) * | 2005-05-16 | 2006-11-20 | Саратовский государственный университет им. Н.Г. Чернышевского (RU) | METHOD FOR TREATING EYE NISTAGMA |
| CN102409095A (en) * | 2011-11-17 | 2012-04-11 | 哈尔滨医科大学 | Congenital nystagmus gene detection kit |
| US20180346981A1 (en) * | 2014-11-20 | 2018-12-06 | Massachusetts Eye And Ear Infirmary | Panel-based Genetic Diagnostic Testing for Inherited Eye Diseases |
| CN108676865A (en) * | 2018-04-08 | 2018-10-19 | 复旦大学附属眼耳鼻喉科医院 | A kind of glaucoma of childhood related gene chip and its preparation method and application |
Non-Patent Citations (2)
| Title |
|---|
| MERVYN G THOMAS等: ""Development and clinical utility of a novel diagnostic nystagmus gene panel using targeted next-generation sequencing"", 《EUROPEAN JOURNAL OF HUMAN GENETICS》 * |
| 赵军等: "利用全外显子组测序法筛选Leber先天性黑一家系候选致病基因的实验研究", 《中华眼科医学杂志(电子版)》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111363804A (en) * | 2020-04-29 | 2020-07-03 | 中信湘雅生殖与遗传专科医院有限公司 | Detection method, detection composition and detection kit for Joubert syndrome |
| CN111363804B (en) * | 2020-04-29 | 2021-08-24 | 中信湘雅生殖与遗传专科医院有限公司 | Detection method, detection composition and detection kit for Joubert syndrome |
| CN111778281A (en) * | 2020-07-17 | 2020-10-16 | 四川省人民医院 | Construction method and application of retina bipolar cytopathy model |
| CN113774128A (en) * | 2021-09-24 | 2021-12-10 | 中国人民解放军东部战区总医院 | Gja8 application of gene mutation site in preparation of product for diagnosing cataract disease |
| CN117778562A (en) * | 2023-04-14 | 2024-03-29 | 湖北省妇幼保健院（湖北省妇女儿童医院） | UBE3B mutant gene, detection kit and method |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN110904209A (en) | DNA library for detecting and diagnosing nystagmus disease-causing gene and application thereof | |
| US20160281166A1 (en) | Methods and systems for screening diseases in subjects | |
| Chaubey et al. | Low-pass genome sequencing: validation and diagnostic utility from 409 clinical cases of low-pass genome sequencing for the detection of copy number variants to replace constitutional microarray | |
| CN106399504A (en) | Targeting-based new generation sequencing deafness gene detection set and kit, and detection method | |
| CN109161591A (en) | Monogenic inheritance kidney trouble gene association screening method, kit and preparation method thereof | |
| CN112813156A (en) | DNA library for detecting and diagnosing bone development disorder pathogenic gene and application thereof | |
| CN109402132B (en) | Nucleic acid for coding SCN1A gene mutant and application thereof | |
| WO2023056451A1 (en) | Compositions and methods for assaying for and genotyping genetic variations | |
| US10720226B2 (en) | Method to match organ donors to recipients for transplantation | |
| CN105648043A (en) | Kit and uses of kit in detection of shortstature related gene | |
| CN112708670A (en) | DNA library for detecting and diagnosing congenital thyroid disease pathogenic gene and application thereof | |
| CN110904128A (en) | Nucleic acid for encoding TEK gene mutant and application thereof | |
| Mei et al. | Clinical and genetic etiologies of neonatal unconjugated hyperbilirubinemia in the China neonatal genomes project | |
| CN110724739A (en) | DNA library for detecting and diagnosing disease-causing genes related to multiple cow's milk coffee spots and application thereof | |
| CN110747270A (en) | DNA library for detecting adult diabetes pathogenic gene for diagnosing juvenile onset and application thereof | |
| CN109402131B (en) | Nucleic acid for encoding KANSL1 gene mutant and application thereof | |
| KR20220024141A (en) | A method for detecting the level of Helicobacter pylori in a fecal sample | |
| Li et al. | Concordance of whole-genome amplified embryonic DNA with the subsequently born child | |
| CN108342404A (en) | INPP5E gene mutation bodies and its application | |
| CN111690738B (en) | Reagent set and kit for detecting POU3F4 gene | |
| CN114164271B (en) | Primer pair, kit and method for detecting breast cancer susceptibility gene variation based on long fragment PCR | |
| CN112877421B (en) | Scar-related biomarker and application thereof | |
| CN114317685B (en) | Kit for detecting mRNA variable shear variation, library building method and sequencing method | |
| WO2023179766A1 (en) | Method for preparing dna library and detecting retroviral integration site | |
| EP4276195A1 (en) | Methods for determining the presence or risk of developing myotonic dystrophy type 1 (md1) or myotonic dystrophy type 2 (md2) |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |