CN105648043A - Kit and uses of kit in detection of shortstature related gene - Google Patents
Kit and uses of kit in detection of shortstature related gene Download PDFInfo
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- CN105648043A CN105648043A CN201410639462.8A CN201410639462A CN105648043A CN 105648043 A CN105648043 A CN 105648043A CN 201410639462 A CN201410639462 A CN 201410639462A CN 105648043 A CN105648043 A CN 105648043A
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Abstract
The present invention provides a kit, uses of the kit in shortstature gene detection, and a method and an apparatus for detecting the shortstature related gene. With the kit, the method and/or the apparatus, the shortstature related gene region can be completely captured in one time, and the variation condition of the shortstature related gene can be detected.
Description
Technical field
The present invention relates to field of gene detection, especially, relate to test kit and the purposes in the short and small genes involved of detection thereof, more particularly, the present invention relates to a kind of test kit, the purposes of test kit in the short and small genes involved of detection, detects the method for short and small genes involved and a kind of device detecting short and small genes involved.
Background technology
(short and small) of short and small stature refers under similar living environment, the more same race of height, same to sex, with more than low 2 standard deviations (-2SD) of age healthy children height average or be in below the 3rd percentile. Short and small is one of common clinical main suit sign of paediatrics, and in crowd, overall incidence is about 3%, and its Causing Factors comprises physiological reasons and pathologic reason. Wherein, pathologic cretinism (cretinism) accounts for short and small overall 30%-40%, its cause of disease comprises that chronic disease, chromosome abnormalty, syndrome, skeleton development are bad, growth hormone deficiency and the insensitive etc. [KantSG of acceptor, WitJM, BreuningMH.Geneticanalysisofshortstature.HormRes.2003; 60 (4): 157-65]. It is the important factor causing cretinism that hereditary defect (comprises single-gene sudden change). Traditional cretinism gene tester relies on accurate clinical diagnosis and method [SeaverLH1, the IronsM of Sanger checking; AmericanCollegeofMedicalGenetics (ACMG) ProfessionalPracticeandGuidelinesCommittee.GenetMed.2009 Jun; 11 (6): 465-70], owing to cretinism has the clinical and genetic heterogeneity of height, the clinical sign of multiple syndrome is overlapping, and traditional cretinism gene tester needs to consume a large amount of time and cost. Therefore, exploitation method efficient, quick, economic, that multiple known short and small Disease-causing gene can carry out abrupt climatic change simultaneously is extremely important to the researchdevelopment of cretinism and clinical diagnosis.
Summary of the invention
According to an aspect of of the present present invention, it is provided that a kind of test kit, it comprises chip, and described chip is fixed on solid-phase matrix by probe to form, and described probe can the exon region of 25 genes below specific recognition: FMR1, BRCC3, WDR81, SATB2, VRK2, LSM14A, HOXD3, LHX1, RIMS1, MBP, HMX2, GJA8, MBD5, ELN, CHRM3, RPS7P5, FMN2, USP9Y, SRY, UTY, CHL1, LBR, RAI1, FGFR3 and RUNX2. The mark that these 25 genes can be used as the micro-deleted micro-repetition of karyomit(e) marks 17 kinds by micro-deleted micro-cretinism repeating to cause of karyomit(e).
Optionally, probe on the chip that test kit comprises or chip can also the exon region of 53 genes below specific recognition: PHEX, ENPP1, FGF23, CLCN5, SLC34A3, VDR, CYP2R1, CYP27B1, SLC37A4, AGL, PFKM, PHKA1, PHKB, PHKA2, ALDOA, PGAM2, G6PC, GALNS, IDUA, IDS, ARSB, HYAL1, GUSB, GLB1, GNPTG, GNPTAB, MAN2B1, MGAT2, SLC35C1, TMEM165, GPD1, GBA, SMPD1, MLYCD, CTSA, UROC1, FUCA1, PIGO, AGA, GK, CTNS, PHGDH, HGD, ATP7A, SLC17A5, MPO, MVK, ADA, PCCB, PLOD3, SLC6A19, ACADS and SUMF1. the single nucleotide variations of these 53 genes can cause 53 kinds of metabolic system diseases relevant to cretinism.
Optionally, the probe on test kit comprises chip or chip can also the exon region of 44 genes below specific recognition: SOX3, GHSR, GHRHR, GHR, BTK, GH1; SHH, SLC29A3, NDN, CEP57, ALMS1, SNRPN, NKX2-5, TSHB, THRA, PAX8, TSHR, IYD, TPO, TG, DUOXA2, DUOX2, SLC26A4, SLC5A5, GCM2, SECISBP2, THRB, TRHR, TRH, PROP1, OTX2, POU1F1, HESX1, LHX4, LHX3, PCNT, RNU4ATAC, IGF1R, GNAS, HSD11B2, B3GALTL, INSR, CYP11B1 and CASR. These 44 genovariations can cause 47 kinds of endocrine system diseases relevant to cretinism.
Optionally, probe on the chip that test kit comprises or chip can also the exon region of 113 genes below specific recognition: TRIP11, COL2A1, ARSE, PTH1R, IMPAD1, EBP, GNPAT, PEX7, AGPS, BMPR1B, RMRP, EXT1, PDE4D, PRKAR1A, GDF5, NPR2, ROR2, NOG, PTHLH, HOXD13, IHH, BMPR1B, HDAC4, SOX9, NEK1, WDR35, DYNC2H1, PIK3R2, AKT3, COL9A3, COL9A2, COL9A1, COMP, MATN3, SLC26A2, MMP13, COL10A1, MMP9, SERPINH1, CRTAP, BMP1, FKBP10, LEPRE1, IFITM5, SERPINF1, PPIB, COL1A1, SP7, MMP13, PAPSS2, MATN3, TRAPPC2, KIF22, ACP5, DDR2, ACAN, COL2A1, CHST3, HES7, DLL3, MCPH1, ASPM, MYH3, TPM2, TNNT3, IFT43, SOST, COL11A1, SEC23A, MMP2, ESCO2, RIN2, LMX1B, GHR, SLC35D1, EFNB1, DYM, LIFR, GORAB, SH3BP2, FGFR1, TBX15, CANT1, TFAP2A, TMCO1, CHRNA1, CTSK, FLNB, EVC, FBN1, B3GAT3, EFTUD2, EIF2AK3, EVC2, CHRNG, CHRND, SMAD4, POC1A, TBX3, WNT7A, ERCC6, ERCC2, FBN1, ADAMTSL2, EXT2, TRPV4, SF3B4, CA2, WDR19, GDF6, FAM123B, TNFRSF11B and TNNI2. these 113 genovariations and aforesaid micro-deleted micro-FGFR3 and RUNX2 gene repeating to cause cretinism, can cause 124 kinds of disease of skeletal system relevant to cretinism.
Optionally, the probe on test kit comprises chip or chip can also the exon region of 5 genes below specific recognition: ATP8B1, ABCB11, BUB1B, CASP8 and CCBE1. These 5 genes can cause 5 kinds of digestive system relevant to cretinism.
Optionally, the probe on test kit comprises chip or chip can also the exon region of 17 genes below specific recognition: ATP6V0A2, LTBP4, PYCR1, ALDH18A1, DKC1, TERT, TINF2, ERCC3, GTF2H5, EXT1, TRPS1, PLEC1, PORCN, SLC39A4, C16orf57, IKBKG and ZMPSTE24.These 17 arbitrary single nucleotide variations of gene can cause the dermatosis relevant to cretinism.
Optionally, the probe on test kit comprises chip or chip can also the exon region of 36 genes below specific recognition: SLC9A6, SMS, HCFC1, NSUN2, RAB40AL, CASK, KDM5C, RPS6KA3, CUL4B, CHMP1A, EXOSC3, AP4S1, AP4E1, AP4B1, GRM1, ZNF592, SEPN1, LARGE, SEPT9, RAPSN, COL6A2, RTTN, SOX3, NAA10, NF1, LRP5, ATM, SHROOM4, SIL1, SLC6A8, IGBP1, SHH, NDE1, IKBKAP, KIF1A and VLDLR. These 36 genes can cause 36 kinds of neuromuscular system diseases relevant to cretinism.
Optionally, the probe on test kit comprises chip or chip can also the exon region of 11 genes below specific recognition: BCOR, STRA6, OTX2, SMOC1, HCCS, SOX2, FOXI1, KCNJ10, SLC26A4, LTBP3 and CLCN5. These 11 genes can cause 11 kinds of ears relevant to cretinism, nose, larynx face systemic disease.
Optionally, the probe on test kit comprises chip or chip can also the exon region of 18 genes below specific recognition: FANCE, FANCC, FANCA, FANCG, FANCF, RAD51C, FANCD2, SLX4, HBB, HBA1, SLC19A2, RPS19, KLF1, GATA1, RARA, ATRX, RBM8A and UROS. These 18 genes add that the variation of the micro-deleted micro-LBR gene repeating to cause cretinism of aforesaid generation can cause 19 kinds of diseases in the blood system relevant to cretinism.
Optionally, the probe on test kit comprises chip or chip can also the exon region of 6 genes below specific recognition: LAMTOR2, LRBA, BTK, WISP3, ITCH and SMARCAL1. The exception of these 6 genes can cause 6 kinds of disease of immune system relevant to cretinism.
Optionally, probe on the chip that test kit comprises or chip can also the exon region of 145 genes below specific recognition: CUL7, CCDC8, KCNJ1, SLC12A2, ERCC8, ERCC6, SMC1A, NIPBL, SMC3, HDAC8, RAD21, ADAMTS2, SLC39A13, COL1A2, B4GALT7, COL3A1, COL1A1, COL5A2, COL5A1, MYCN, MIR17HG, ORC6, ORC1, CDT1, CDC6, ORC4, KRAS, RAF1, BRAF, SOS1, PTPN11, OFD1, TCTN3, WNT5A, ROR3, CREBBP, EP300, LIFR, HSPG2, ATR, RBBP8, CEP152, NIN, CEP63, CENPJ, ADAMTS17, FBN1, LTBP2, ADAMTS10, RAB3GAP1, RAB3GAP2, RAB18, FOXG1, MECP2, LRP2, NKX2-1, FGD1, KCNJ2, SBDS, L1CAM, AAAS, TWIST1, NBAS, SEMA3E, PQBP1, SPG20, G6PC3, HYLS1, NBN, IGF2, UBR1, OCRL, CHD7, GLA, CD96, LMNA, PHF6, HRAS, IFT140, SRCAP, CTC1, OTX2, ASXL1, FTO, ANKRD11, HSD17B4, MED12, KIF7, DHCR7, CTDP1, MYH8, DDX11, POR, KAT6B, ALDH3A2, TP63, SDHA, ALPL, MLL, WRN, MAP2K1, NSDHL, FAM20C, NSD1, H19, RECQL4, TRIM37, ZEB2, FOXE1, KANSL1, BANF1, BLM, VPS13B, RAB23, TBX1, HPRT1, SLC16A2, SMARCA2, COX4I2, MAP2K2, NOTCH2, CRLF1, WFS1, ZBTB16, ARX, DLX5, MGP, FGFR2, TAZ, PITX2, ACTB, PLOD2, SLC34A1, MBTPS2, TMEM237, TBCE, MKS1, GPC6, RIPK4, FLNA, GJB6, ERCC3, MPV17, ATPAF2 and AQP2.The exception of these 145 genes and the micro-deleted micro-BAI1 gene repeating to cause cretinism of aforesaid generation can cause 153 kinds of syndromes relevant to cretinism.
Optionally, the probe on described test kit comprises chip or chip can the exon region of 438 genes below specific recognition: AAAS, ABCB11, ACADS, ACAN, ACP5, ACTB, ADA, ADAMTS10, ADAMTS17, ADAMTS2, ADAMTSL2, AGA, AGL, AGPS, AKT3, ALDH18A1, ALDH3A2, ALDOA, ALMS1, ALPL, FAM123B, ANKRD11, AP4B1, AP4E1, AP4S1, AQP2, ARSB, ARSE, ARX, ASPM, ASXL1, ATM, ATP6V0A2, ATP7A, ATP8B1, ATPAF2, ATR, ATRX, AVPR2, B3GALTL, B3GAT3, B4GALT7, BANF1, BCOR, BLM, BMP1, BMPR1B, BRAF, BTK, BUB1B, CA2, CANT1, CASK, CASP8, CASR, CCBE1, CCDC8, CD96, CDC6, CDSN, CDT1, CENPJ, CEP152, CEP57, CEP63, CHD7, CHMP1A, CHRNA1, CHRND, CHRNG, CHST3, CLCN5, COL10A1, COL11A1, COL1A1, COL1A2, COL2A1, COL3A1, COL5A1, COL5A2, COL6A2, COL9A1, COL9A2, COL9A3, COLEC11, COMP, COX4I2, CREBBP, CRLF1, CRTAP, CTC1, CTDP1, CTNS, CTSA, CTSK, CUL4B, CUL7, CYP11B1, CYP27B1, CYP2R1, DDR2, DDX11, DHCR7, DHODH, DKC1, DLL3, DLX5, DUOX2, DUOXA2, DYM, DYNC2H1, EBP, EFNB1, EFTUD2, EIF2AK3, ENPP1, EP300, ERCC2, ERCC3, ERCC6, ERCC8, ESCO2, EVC, EVC2, EXOSC3, EXT1, EXT2, FAM20C, FANCA, FANCC, FANCD2, FANCE, FANCF, FANCG, FBN1, FGD1, FGF23, FGFR1, FGFR2, FKBP10, FLNA, FLNB, FOXE1, FOXG1, FOXI1, FTO, FUCA1, G6PC, G6PC3, GALNS, GATA1, GBA, GCM2, GDF5, GDF6, GH1, GHR, GHRHR, GHSR, GJB6, GK, GLA, GLB1, GLI2, GNAS, GNPAT, GNPTAB, GNPTG, GORAB, GPC6, GPD1, GRM1, GTF2H5, GUSB, GYS2, H19, HBA1, HBA2, HBB, HCCS, HCFC1, HDAC4, HDAC8, HES7, HESX1, HGD, HOXD13, HPRT1, HRAS, HSD11B2, HSD17B4, HSPG2, HYAL1, HYLS1, IDS, IDUA, IFITM5, IFT140, IFT43, IGBP1, IGF1, IGF1R, IGF2, IHH, IKBKAP, IKBKG, IMPAD1, INSR, ITCH, IYD, KANSL1, KAT6B, KCNJ1, KCNJ10, KCNJ2, KDM5C, KDM6A, KIF1A, KIF22, KIF7, KLF1, MLL, MLL2, KRAS, L1CAM, LAMTOR2, LARGE, LEPRE1, LHX3, LHX4, LIFR, LMNA, LMX1B, LRBA, LRP2, LRP5, LTBP2, LTBP3, LTBP4, MAN2B1, MAP2K1, MAP2K2, MASP1, MATN3, MBTPS2, MCPH1, MECP2, MED12, MGAT2, MGP, MIR17HG, MKS1, MLYCD, MMP13, MMP2, MMP9, MPO, MPV17, MVK, MYCN, MYH3, MYH8, NAA10, NBAS, NBN, NDE1, NDN, NEK1, NF1, NIN, NIPBL, NKX2-1, NKX2-5, NOG, NOTCH2, NPR2, NSD1, NSDHL, NSUN2, OCRL, OFD1, ORC1, ORC4, ORC6, OTX2, PAPSS2, PAX8, PCCB, PCNT, PDE4D, PEX7, PFKM, PGAM2, PHEX, PHF6, PHGDH, PHKA1, PHKA2, PHKB, PIGO, PIK3R2, PITX2, PLEC, PLOD2, PLOD3, POC1A, POR, PORCN, POU1F1, PPIB, PQBP1, PRKAR1A, PROP1, PTH1R, PTHLH, PTPN11, PYCR1, RAB23, RAB3GAP1, RAB3GAP2, RAB40AL, RAD21, RAD51C, RAF1, RAPSN, RARA, RBBP8, RBM8A, RECQL4, RIN2, RIPK4, RMRP, RNU4ATAC, ROR2, RPS19, RPS6KA3, RTTN, SBDS, SDHA, SEC23A, SECISBP2, SEMA3E, SEPN1, SEPT9, SERPINF1, SERPINH1, SF3B4, SH3BP2, SHH, SHROOM4, SIL1, SLC12A1, SLC16A2, SLC17A5, SLC19A2, SLC26A2, SLC26A4, SLC29A3, SLC34A1, SLC34A3, SLC35C1, SLC35D1, SLC37A4, SLC39A13, SLC39A4, SLC5A5, SLC6A19, SLC6A8, SLC9A6, SLX4, SMAD4, SMARCA2, SMARCAL1, SMC1A, SMC3, SMOC1, SMPD1, SMS, SNRPN, SOS1, SOST, SOX2, SOX3, SOX9, SP7, SPG20, SRCAP, STRA6, SUMF1, TAZ, TBCE, TBX1, TBX15, TBX3, TCTN3, TERT, TFAP2A, TG, THRA, THRB, TINF2, TMCO1, TMEM165, TMEM237, TNFRSF11B, TNNI2, TNNT3, TP63, TPM2, TPO, TRAPPC2, TRH, TRHR, TRIM37, TRIP11, TRPS1, TRPV4, TSHB, TSHR, TWIST1, UBR1, UROC1, UROS, USB1, VDR, VLDLR, VPS13B, WDR19, WDR35, WFS1, WISP3, WNT5A, WNT7A, WRN, ZBTB16, ZEB2, ZMPSTE24 and ZNF592.This 438 single-genes sudden change can cause 471 kinds of single-gene disorders relevant to cretinism, add aforementioned choose in order to mark 17 kinds by 25 genes of micro-deleted micro-cretinism repeating to cause of karyomit(e). these 463 genes are that contriver obtains through collection, repeatedly screening and composite test, it is possible to represent comprehensively or represent cretinism genes involved. the disease that these 463 genes are relevant to 488 kinds of cretinisms is correlated with, with reference to aforementioned, in 463 genes, the SNP variation of 53 genes can cause 53 kinds of metabolic system diseases relevant to cretinism, the SNP occurred in 44 genes can cause 47 kinds of endocrine system diseases relevant to cretinism, the SNP of 115 genes can cause 124 kinds of disease of skeletal system relevant to cretinism, the SNP of 17 genes can cause 17 kinds of dermatosis relevant to cretinism, 146 genes can cause 153 kinds of syndromes relevant to cretinism, 36 genes can cause 36 kinds of neuromuscular system diseases relevant to cretinism, 19 genes can cause 19 kinds of diseases in the blood system relevant to cretinism, 11 genes can cause 11 kinds of ears relevant to cretinism, nose, larynx face systemic disease, 6 genes can cause 6 kinds of disease of immune system relevant to cretinism, 5 genes can cause 5 kinds of digestive system relevant to cretinism.
Optionally, the probe on described test kit comprises chip or chip can also specific recognition above-mentioned its can the intron region of each 30bp of upstream and downstream of exon region of each gene of specific recognition. Specificity and efficiency when being conducive to raising chip or probe to catch like this.
According to the another aspect of the present invention, also provide the purposes of mentioned reagent box in the short and small genes involved of detection. Detect short and small genes involved time, test kit can catch short and small genes involved region in order to specific recognition, with obtain target area related data information.
According to another aspect of the invention, it is provided that a kind of method detecting short and small genes involved, described method comprises: (1) obtains the nucleic acid of person under inspection, and described nucleic acid is genomic nucleic acids and/or free nucleic acid fragment, (2) catch described nucleic acid, obtain short and small genes involved region, (3) described short and small genes involved region is carried out sequencing, obtain sequence information, (4) described short and small genes involved is detected based on described sequence information, wherein, (2) utilize the arbitrary test kit in one aspect of the present invention and each enforcement mode to carry out. the aforementioned advantage about the test kit of one aspect of the present invention or the purposes of test kit and technology feature, be suitable for the method for this one side of the present invention equally. in one embodiment of the invention, (4) comprise the SNP and micro-deleted micro-repetition (INDEL) variation that detect described short and small genes involved based on described sequence information simultaneously, specifically, detect these two kinds of variations to comprise: described sequence information and reference sequences are carried out the first comparison, obtains the first comparison result simultaneously, a part for described first comparison result and described reference sequences is carried out the 2nd comparison, obtains the 2nd comparison result, based on described first comparison and described 2nd comparison result, the SNP and micro-deleted micro-repetition that detect described short and small genes involved make a variation simultaneously. here, said first comparison is overall comparison, 2nd comparison is local comparison, first comparison is conventional comparison, can utilize but be not limited to the softwares such as SOAP or BWA and carry out according to its default setting, obtain the first comparison result, first comparison result comprises the matched position of sequence information on reference sequences and match condition information, in one embodiment of the invention, said reference sequences is hg19, in 2nd comparison, a part for said reference sequences comprises each the known INDEL site in the reference sequences corresponding with the short and small genes involved region caught, and the reference sequences of each known INDEL site each 1000bp of upstream and downstream described, carry out the 2nd comparison and namely carry out local comparison again based on all sequences information (reads) near all known INDEL in the result pair of the first comparison reference sequences corresponding with the short and small genes involved region caught, the mistake in the first comparison can be eliminated, improve the accuracy rate of follow-up variation detection, 2nd comparison can utilize GATK anharmonic ratio to software (https://www.broadinstitute.org/gatk/) carry out.In one embodiment of the invention, detect described SNP and INDEL to make a variation simultaneously, being undertaken by GATKUnifiedGenotyper software, the INDEL gone out according to this software detection has more false positive, utilizes the 2nd comparison above i.e. local anharmonic ratio to reduce false positive INDEL to being conducive to. The method of this one side of the present invention is that based target areas captured is in conjunction with s-generation high throughput sequencing technologies, order-checking can be caught in the coding region of little 463 the relevant genes of short disease and flank �� 30bp region, then these genovariations are carried out determination and analysis, it is a kind of efficient, quick, economic cretinism gene tester. The mutational range that can detect comprises point mutation and small segment insertion and deletion. The cretinism gene tester based on the order-checking of two generations of the present invention, by the abnormal variation of order-checking detection is carried out analyze, specify the pathogenic mutation of person under inspection, it is achieved to the accurate diagnosis of cretinism disease, treat and prevent disease from recurring in family targetedly.
According to the one side again of the present invention, a kind of device detecting short and small genes involved is provided, this device can in order to perform the part Overall Steps alive realizing the method for aforementioned the present invention, this device comprises: A. nucleic acid acquiring unit, for obtaining the nucleic acid of person under inspection, described nucleic acid is genomic nucleic acids and/or free nucleic acid fragment; B. capturing unit, is connected with A unit, for catching from the nucleic acid in A unit, to obtain short and small genes involved region; C. sequencing unit, is connected with unit B, for the short and small genes involved region from unit B is carried out sequencing, to obtain sequence information; D. detecting unit, is connected with C cell, for based on the sequence information described short and small genes involved of detection from C cell; Wherein, catching in unit B utilizes the arbitrary chip in aforementioned one aspect of the present invention and each enforcement mode to carry out. Fig. 1 is the apparatus structure schematic diagram in one embodiment of the present of invention. To the advantage of method and the description of technology feature of the present invention, the same device being suitable for the present invention, and, those skilled in the art are appreciated that, all or part of unit in the device of the present invention, selectable, dismountable comprises one or more subelement to perform or to realize each embodiment of aforementioned the inventive method.
Accompanying drawing explanation
Above-mentioned and/or the additional aspect of the present invention and advantage from accompanying drawing below combining to the description of the mode of enforcement becoming obviously with it should be readily understood that wherein:
Fig. 1 is the schematic diagram of the device of the short and small genes involved of detection in a specific embodiment of the present invention;
Fig. 2 is the schema of the short and small genes involved of detection in a specific embodiment of the present invention.
Embodiment
Gene title in the application's description all adopts the official in NCBI-Gene to name (OfficialSymbol). In addition, the missense mutation (missense) occurred in description refers to, encodes certain amino acid whose codon after base is replaced, becomes the codon of another seed amino acid of coding, thus the amino acid kind of polypeptide chain and sequence are changed. Some missense mutation can make polypeptide chain lose original function, and the exception of numerous protein is caused by missense mutation.
According to general method, it is achieved the method for the present invention mainly comprises the design of short and small genes involved detection chip, and the exploitation of sequencing technologies and analysis process is caught in target area.
(1) cretinism gene detecting chip design
Chromosomal variation and single-gene can cause cretinism, and chromosomal variation comprises again the change of chromosome number, structure, and single parent diploid and chimerism can also cause cretinism. below only example utilize the inventive method detection single-gene and part karyomit(e) micro-deleted and micro-repeat to cause of short and small stature. by searching omim database and pertinent literature, obtain 471 kinds of single-gene disorders relevant to cretinism that 438 single-genes cause, have chosen 25 genes and mark 17 kinds by micro-deleted micro-cretinism repeating to cause of karyomit(e) as the marker of the micro-deleted micro-repetition of karyomit(e). wherein, the SNP variation of 53 genes can cause 53 kinds of metabolic system diseases relevant to cretinism, the SNP occurred in 44 genes can cause 47 kinds of endocrine system diseases relevant to cretinism, the SNP of 115 genes can cause 124 kinds of disease of skeletal system relevant to cretinism, the SNP of 17 genes can cause 17 kinds of dermatosis relevant to cretinism, 36 genes can cause 36 kinds of neuromuscular system diseases relevant to cretinism, 19 genes can cause 19 kinds of diseases in the blood system relevant to cretinism, 11 genes can cause 11 kinds of ears relevant to cretinism, nose, larynx face systemic disease, 6 genes can cause 6 kinds of disease of immune system relevant to cretinism, 5 genes can cause 5 kinds of digestive system relevant to cretinism, and 146 genes can cause 153 kinds of other syndromes relevant to cretinism. there will be 1 gene in these genes and cause multiple disease, like this, the gene chosen amounts to 463, corresponding 488 kinds of diseases, specifically as shown in table 1, the gene that in table 1, the gene representation of runic display is relevant to multiclass disease, table 2 shows number gene and corresponding disease information of number.
According to human genome HG19, exon sequence and the flank �� 30bp region of choosing above-mentioned 463 genes carry out probe chip design, can designing probe sequence like this: from hg19, obtain the exon sequence of above-mentioned 463 genes and each section of reference sequences in flank �� 30bp region, to each section of reference sequences, all from one end of one section of reference sequences, the reference sequences copying predetermined length successively obtains probe sequence, make finally total probe can cover this section of reference sequences at least one times, can be overlapping or not overlapping between adjacent probe sequence, here, predetermined length is the length of probe, can be 50-250nt, then chip is prepared according to synthesizer specification sheets or trust producer synthesising probing needle. the die size obtained is about 2.5M, this chip covers abundant capture probe, probe covers region and reaches 98.83%, it is possible to enrichment target DNA fragments from the genome of complexity, catches, on same chip, the genome area being about 2.5M with high specific and high coverage rate.
Table 1
Table 2
(2) order-checking and analysis process are caught in target area
As shown in Fig. 2 test and analysis process, from person under inspection's whole blood, extract genomic dna, and the qualified DNA of detection is carried out SNP mass spectrometric detection and library preparation simultaneously. library preparation is that 1 �� g genomic dna is broken into master tape is 200-300bp small pieces segment DNA, then will interrupt rear DNA fragmentation and carry out end-filling, base " A " is added at 3' end, make DNA fragmentation that the special joint of " T " base can be had to be connected with 3' end band, through the library that Non-CapturedPCR (not catching front amplification) has built, by cretinism gene detecting chip by the Exon of the specific gene chosen and and flank �� 30bp region carry out enrichment, again by pcr amplification enrichment after product, sequence capturing hybridization efficiency is obtained finally by PCR primer QPCR detection before and after hybridization.After QPCR detection is qualified, the library of some amount is carried out upper machine order-checking, such as, utilizes Hiseq2000/2500, lower machine data is carried out Quality Control, then data are analyzed and understand. Wherein, a sample cycle is prepared in library is 5-7 days. Information analysis adopts the information analysis flow process of independent development to carry out data processing, software that what analysis process comprised do not explain especially, script etc. obtain by webpage or database disclosed in the Hua Da gene of Shenzhen, or the service of customization Shenzhen Hua Da gene carries out this data processing, data processing mainly comprises filtration, comparison, deduplication, anharmonic ratio to steps such as, Quality Control, SNV (SNP)+INDEL detection, annotations. The deciphering of annotation result, mainly consult based on HGMD, BGIGap and each big deaf pathogenic mutation database and literature search and carry out, comprehensively understand in conjunction with multiple function prediction software results and the clinical sign of person under inspection simultaneously, primitive rule carries out with reference to U.S.'s medicogenetics and genomics institute (AmericanCollegeofMedicalGeneticsandGenomics, ACMG) associated guideline.
Traditional cretinism gene tester relies on the accurate diagnosis of clinician, by the Sanger of candidate gene verifies the pathogenesis seeking patient. If the Sanger of candidate gene is verified as feminine gender, it is necessary to again clinical sign is assessed and find new candidate gene, waste a large amount of time and cost. The method of the present invention of example based on two generation sequencing technologies cretinism gene test can select gene set flexibly, and cost performance is higher, specific aim is stronger, is applicable to large-scale clinical gene test service. The cretinism gene test that present method provides covers the cretinism that the single-gene reported at present causes substantially, it is possible to carry out person under inspection accurately diagnosing, treating targetedly and prevent disease from recurring in family. And, along with the discovery of the new genes involved of cretinism, this chip can constantly be upgraded, and adds new gene, thus constantly promotes the recall rate of cretinism chip.
Below in conjunction with concrete individual specimen, the operation result of the concrete detection method according to the present invention is described in detail. Example below, only for explaining the present invention, and can not be interpreted as limitation of the present invention. In describing the invention, unless otherwise explanation, the implication of " multiple " is two or more.
Except as otherwise explaining, the reagent do not explained especially, sequence (joint, label and primer), software and the instrument related in following examples, be all conventional commercially available prod or disclosed in, such as build storehouse related kit to carry out library construction etc. purchased from the hiseq2000 of the Illumina company platform that checks order.
Embodiment
The 1 non-well-balanced property of example is short and small, and skeletal abnormality (convex after congenital vertebral, four limbs are short and small), face feature abnormalities, congenital hepatic artery portal vein fistula, the patient of congenital hydrocephalus detects, and patient is from Tianjin Association for the Handicapped. Which kind of disease this patient cannot make a definite diagnosis as clinically, through above-mentioned in conjunction with cretinism genes involved chip a new generation target area catch order-checking (NGSPanel) detection after, find with the FGFR3 gene of dyschondroplasia on found that the harmful sudden change of a focus is c.1138G > A (p.Gly380Arg), final patient makes a definite diagnosis as dyschondroplasia.
Extracting specimen dna with salting-out process, large fragment DNA carries out ultrasonic interrupting, and uses sample to interrupt method at present and interrupts method for Covaris, sample DNA is smashed the fragment to 100-700bp scope.(note: interrupt effect generally ideal in 200-250bp position with Insert fragment master tape position, required preparation library, if it is undesirable to interrupt effect, need again to interrupt. )
1. library preparation
1.1 end reparation and purifying
By mixed for the mix concussion that configures even after, each reaction adds 25 �� L enzyme reaction mixed solutions. Reaction conditions: 20 DEG C, 30min.
Using 180 �� LAmpureBeads to carry out product purification, the DNA of recovery is dissolved in the water of 30 �� L (wherein 1.9 �� L are loss).
1.2 ends add " A " (A-Tailing)
By mixed for the mix concussion that configures even after, often pipe adds 6.9 �� L enzyme reaction mixed solutions. Reaction conditions: 20 DEG C, 30min. It should be noted that, end can without purifying after adding " A ".
The connection of 1.3Adapter and purifying
Mixing even by the mix concussion configured, each reaction adds 15 �� L enzyme reaction mixed solutions. Reaction conditions: 16 DEG C, 12-16h (spends the night). Using 75 �� LAmpureBeads to carry out product purification, the DNA of recovery is dissolved in the water of 35 �� L (wherein 2 �� L are loss).
1.4Non-Captured sample P re-LM-PCR
PCR program:
94 DEG C of 2min;
94 DEG C of 15s, 62 DEG C of 30s, 72 DEG C of 30s, 4cycles;
72 DEG C of 5min;
4��forever
2. chip hybridization, enrichment is caught in target area
This experiment carries out hybridization elution with reference to NimbleGen working instructions, obtains goal gene and PCR enrichment.
3. go up machine order-checking
This experiment adopts hiseq2000 or hiseq2500PE101+8+101 program to carry out upper machine order-checking.
4. information analysis
4.1 obtain raw data (FASTQ data) from sequenator.
4.2 filter: original FASTQ data are carried out quality control, removes conventional said low quality value data.
4.3 comparisons: utilize SOAP software and default parameters thereof to arrange, it may also be useful to Hg19 reference sequences is compared, parallelization Processing tasks.
4.4 deduplications: based on the read deduplication algorithm of Picard, find out repetition reads from comparison result to parallelization and mark in the tag mode of SAM/BAM file.
4.5 anharmonic ratioes pair: use based on the heavy comparison model of GATK, carry out anharmonic ratio pair. That is, on the basis of a upper comparison result, all reads near INDEL are carried out local comparison again, to eliminate the mistake of comparison, it is to increase the accuracy rate of variation detection.
4.6 detection SNVINDEL: be used on GATKUnifiedGenotyper basis exploitation the parallelization based on Hadoop platform variation detection module, carry out the detection of SNP and Indel simultaneously. Than to utilizing after completing more complete wrong model to calculate genotype likelihood value, it is contemplated that multiple factors such as PCR, base quality (BaseQuality) and comparison quality (MappingQuality). A kind of variation that UnifiedGenotyper is the multiple mutation detection method of set and becomes identifies software (VariantsCaller), UnifiedGenotyper uses Bayes's Maximum Likelihood Model, estimating genotype and gene frequency, finally each variant sites and genotype to each sample can provide an accurate posterior probability simultaneously. Sample in input bam file can be carried out variation detection by this software, finally generates a vcf file, can comprise variant sites and the genotype information of all samples in vcf file. The result obtained from vcf file be the most original, not through the Variants set of any filtration and correction. The variant sites that this step produces has very high false positive, especially indel, and therefore, we carry out anharmonic ratio to the false positive results to reduce in this abrupt climatic change as far as possible above.
4.7 annotations: use human genome database NCBI104, frequency database dbSNP135,1000human, ESP6500, and BGI internal frequency database annotates; Using HGVS that variation is carried out standard name, use OMIM, HGMD disease database simultaneously, the clinical genome database of CGD carries out suddenling change and disease annotation, it is determined that gene that mutational site occurs, coordinate, amino acid change etc. Generate the accidental data list relevant to cretinism gene.
5. result analysis
Table 3 is sequencing data statistics, and table 4 display-object area coverage Statistical information, table 5 shows detected result.
Table 3
Table 4
Table 5
As shown in table 5, this gene test with dyschondroplasia (Achondroplasia, ACH; OMIM:100800) relevant FGFR3 gene coding region detects that a known pathogenic mutation of assorted conjunction is c.1138G > A (p.Gly380Arg). The assorted conjunction missense mutation of FGFR3 gene coding region is c.1138G > A (p.Gly380Arg) is the pathogenic mutation of the most common dyschondroplasia, the dyschondroplasia patient of 99% is caused by the sudden change of FGFR3 gene. The nineteen ninety-fives such as Bellus are to finding in the research of 193 example dyschondroplasia patients that 187 example patients are by suddenling change c.1138G > A causes, account for the 96.9% of patient, by another sudden change of this position c.1138G 5 example patients are > C causes, account for the 2.6% [Bellus of patient, G.A., Hefferon, etal.AchondroplasiaisdefinedbyrecurrentG380RmutationsofF GFR3.Am.J.Hum.Genet.1995 (56): p.368-373]. Dyschondroplasia is autosomal dominant inherited disease, and heterozygous mutant can cause a disease. Therefore, c.1138G > A (p.Gly380Arg) should be the reason of person under inspection's skeletal abnormality associated conditions.
In addition, as space is limited, list the information of other variation of this part detected out, as shown in table 6, variation in table 6 is all database medium frequency < 5%, and may affect the variation type (comprising the variation within non-same sense mutation, the insertion of protein-coding region, deletion mutantion and montage �� 10bp) of protein function and mRNA montage.
Table 6
* hom/het:hom represents that this mutational site is homozygous mutation, and het represents that this mutational site is heterozygous mutant.
* the frequency information about this SNP included in Fr.1:dbSNP database.
* Fr.2: thousand people in planning all in order-checking sample about the frequency information of this SNP in asian ancestry's ethnic group.
* the frequency information about this SNP included in Fr.3:ESP6500 database.
* Fr.4: about the frequency information of this SNP in local data base.
* Condel:Condel database predicts the outcome.
Claims (12)
1. a test kit, it comprises chip, and described chip is fixed on solid-phase matrix by probe to form, and described probe can the exon region of 25 genes below specific recognition: FMR1, BRCC3, WDR81, SATB2, VRK2, LSM14A, HOXD3, LHX1, RIMS1, MBP, HMX2, GJA8, MBD5, ELN, CHRM3, RPS7P5, FMN2, USP9Y, SRY, UTY, CHL1, LBR, RAI1, FGFR3 and RUNX2;
Optionally, described probe can the exon region of 53 genes below specific recognition: PHEX, ENPP1, FGF23, CLCN5, SLC34A3, VDR, CYP2R1, CYP27B1, SLC37A4, AGL, PFKM, PHKA1, PHKB, PHKA2, ALDOA, PGAM2, G6PC, GALNS, IDUA, IDS, ARSB, HYAL1, GUSB, GLB1, GNPTG, GNPTAB, MAN2B1, MGAT2, SLC35C1, TMEM165, GPD1, GBA, SMPD1, MLYCD, CTSA, UROC1, FUCA1, PIGO, AGA, GK, CTNS, PHGDH, HGD, ATP7A, SLC17A5, MPO, MVK, ADA, PCCB, PLOD3, SLC6A19, ACADS and SUMF1,
Optionally, described probe can the exon region of 44 genes below specific recognition: SOX3, GHSR, GHRHR, GHR, BTK, GH1; SHH, SLC29A3, NDN, CEP57, ALMS1, SNRPN, NKX2-5, TSHB, THRA, PAX8, TSHR, IYD, TPO, TG, DUOXA2, DUOX2, SLC26A4, SLC5A5, GCM2, SECISBP2, THRB, TRHR, TRH, PROP1, OTX2, POU1F1, HESX1, LHX4, LHX3, PCNT, RNU4ATAC, IGF1R, GNAS, HSD11B2, B3GALTL, INSR, CYP11B1 and CASR;
Optionally, described probe can the exon region of 113 genes below specific recognition: TRIP11, COL2A1, ARSE, PTH1R, IMPAD1, EBP, GNPAT, PEX7, AGPS, BMPR1B, RMRP, EXT1, PDE4D, PRKAR1A, GDF5, NPR2, ROR2, NOG, PTHLH, HOXD13, IHH, BMPR1B, HDAC4, SOX9, NEK1, WDR35, DYNC2H1, PIK3R2, AKT3, COL9A3, COL9A2, COL9A1, COMP, MATN3, SLC26A2, MMP13, COL10A1, MMP9, SERPINH1, CRTAP, BMP1, FKBP10, LEPRE1, IFITM5, SERPINF1, PPIB, COL1A1, SP7, MMP13, PAPSS2, MATN3, TRAPPC2, KIF22, ACP5, DDR2, ACAN, COL2A1, CHST3, HES7, DLL3, MCPH1, ASPM, MYH3, TPM2, TNNT3, IFT43, SOST, COL11A1, SEC23A, MMP2, ESCO2, RIN2, LMX1B, GHR, SLC35D1, EFNB1, DYM, LIFR, GORAB, SH3BP2, FGFR1, TBX15, CANT1, TFAP2A, TMCO1, CHRNA1, CTSK, FLNB, EVC, FBN1, B3GAT3, EFTUD2, EIF2AK3, EVC2, CHRNG, CHRND, SMAD4, POC1A, TBX3, WNT7A, ERCC6, ERCC2, FBN1, ADAMTSL2, EXT2, TRPV4, SF3B4, CA2, WDR19, GDF6, FAM123B, TNFRSF11B and TNNI2,
Optionally, described probe can the exon region of 5 genes below specific recognition: ATP8B1, ABCB11, BUB1B, CASP8 and CCBE1;
Optionally, described probe can the exon region of 17 genes below specific recognition: ATP6V0A2, LTBP4, PYCR1, ALDH18A1, DKC1, TERT, TINF2, ERCC3, GTF2H5, EXT1, TRPS1, PLEC1, PORCN, SLC39A4, C16orf57, IKBKG and ZMPSTE24;
Optionally, described probe can the exon region of 36 genes below specific recognition: SLC9A6, SMS, HCFC1, NSUN2, RAB40AL, CASK, KDM5C, RPS6KA3, CUL4B, CHMP1A, EXOSC3, AP4S1, AP4E1, AP4B1, GRM1, ZNF592, SEPN1, LARGE, SEPT9, RAPSN, COL6A2, RTTN, SOX3, NAA10, NF1, LRP5, ATM, SHROOM4, SIL1, SLC6A8, IGBP1, SHH, NDE1, IKBKAP, KIF1A and VLDLR;
Optionally, described probe can the exon region of 11 genes below specific recognition: BCOR, STRA6, OTX2, SMOC1, HCCS, SOX2, FOXI1, KCNJ10, SLC26A4, LTBP3 and CLCN5;
Optionally, described probe can the exon region of 18 genes below specific recognition: FANCE, FANCC, FANCA, FANCG, FANCF, RAD51C, FANCD2, SLX4, HBB, HBA1, SLC19A2, RPS19, KLF1, GATA1, RARA, ATRX, RBM8A and UROS;
Optionally, described probe can the exon region of 6 genes below specific recognition: LAMTOR2, LRBA, BTK, WISP3, ITCH and SMARCAL1;
Optionally, described probe can the exon region of 145 genes below specific recognition: CUL7, CCDC8, KCNJ1, SLC12A2, ERCC8, ERCC6, SMC1A, NIPBL, SMC3, HDAC8, RAD21, ADAMTS2, SLC39A13, COL1A2, B4GALT7, COL3A1, COL1A1, COL5A2, COL5A1, MYCN, MIR17HG, ORC6, ORC1, CDT1, CDC6, ORC4, KRAS, RAF1, BRAF, SOS1, PTPN11, OFD1, TCTN3, WNT5A, ROR3, CREBBP, EP300, LIFR, HSPG2, ATR, RBBP8, CEP152, NIN, CEP63, CENPJ, ADAMTS17, FBN1, LTBP2, ADAMTS10, RAB3GAP1, RAB3GAP2, RAB18, FOXG1, MECP2, LRP2, NKX2-1, FGD1, KCNJ2, SBDS, L1CAM, AAAS, TWIST1, NBAS, SEMA3E, PQBP1, SPG20, G6PC3, HYLS1, NBN, IGF2, UBR1, OCRL, CHD7, GLA, CD96, LMNA, PHF6, HRAS, IFT140, SRCAP, CTC1, OTX2, ASXL1, FTO, ANKRD11, HSD17B4, MED12, KIF7, DHCR7, CTDP1, MYH8, DDX11, POR, KAT6B, ALDH3A2, TP63, SDHA, ALPL, MLL, WRN, MAP2K1, NSDHL, FAM20C, NSD1, H19, RECQL4, TRIM37, ZEB2, FOXE1, KANSL1, BANF1, BLM, VPS13B, RAB23, TBX1, HPRT1, SLC16A2, SMARCA2, COX4I2, MAP2K2, NOTCH2, CRLF1, WFS1, ZBTB16, ARX, DLX5, MGP, FGFR2, TAZ, PITX2, ACTB, PLOD2, SLC34A1, MBTPS2, TMEM237, TBCE, MKS1, GPC6, RIPK4, FLNA, GJB6, ERCC3, MPV17, ATPAF2 and AQP2.
2. the test kit of claim 1, it is characterised in that, described probe can the exon region of 438 genes below specific recognition: AAAS, ABCB11, ACADS, ACAN, ACP5, ACTB, ADA, ADAMTS10, ADAMTS17, ADAMTS2, ADAMTSL2, AGA, AGL, AGPS, AKT3, ALDH18A1, ALDH3A2, ALDOA, ALMS1, ALPL, FAM123B, ANKRD11, AP4B1, AP4E1, AP4S1, AQP2, ARSB, ARSE, ARX, ASPM, ASXL1, ATM, ATP6V0A2, ATP7A, ATP8B1, ATPAF2, ATR, ATRX, AVPR2, B3GALTL, B3GAT3, B4GALT7, BANF1, BCOR, BLM, BMP1, BMPR1B, BRAF, BTK, BUB1B, CA2, CANT1, CASK, CASP8, CASR, CCBE1, CCDC8, CD96, CDC6, CDSN, CDT1, CENPJ, CEP152, CEP57, CEP63, CHD7, CHMP1A, CHRNA1, CHRND, CHRNG, CHST3, CLCN5, COL10A1, COL11A1, COL1A1, COL1A2, COL2A1, COL3A1, COL5A1, COL5A2, COL6A2, COL9A1, COL9A2, COL9A3, COLEC11, COMP, COX4I2, CREBBP, CRLF1, CRTAP, CTC1, CTDP1, CTNS, CTSA, CTSK, CUL4B, CUL7, CYP11B1, CYP27B1, CYP2R1, DDR2, DDX11, DHCR7, DHODH, DKC1, DLL3, DLX5, DUOX2, DUOXA2, DYM, DYNC2H1, EBP, EFNB1, EFTUD2, EIF2AK3, ENPP1, EP300, ERCC2, ERCC3, ERCC6, ERCC8, ESCO2, EVC, EVC2, EXOSC3, EXT1, EXT2, FAM20C, FANCA, FANCC, FANCD2, FANCE, FANCF, FANCG, FBN1, FGD1, FGF23, FGFR1, FGFR2, FKBP10, FLNA, FLNB, FOXE1, FOXG1, FOXI1, FTO, FUCA1, G6PC, G6PC3, GALNS, GATA1, GBA, GCM2, GDF5, GDF6, GH1, GHR, GHRHR, GHSR, GJB6, GK, GLA, GLB1, GLI2, GNAS, GNPAT, GNPTAB, GNPTG, GORAB, GPC6, GPD1, GRM1, GTF2H5, GUSB, GYS2, H19, HBA1, HBA2, HBB, HCCS, HCFC1, HDAC4, HDAC8, HES7, HESX1, HGD, HOXD13, HPRT1, HRAS, HSD11B2, HSD17B4, HSPG2, HYAL1, HYLS1, IDS, IDUA, IFITM5, IFT140, IFT43, IGBP1, IGF1, IGF1R, IGF2, IHH, IKBKAP, IKBKG, IMPAD1, INSR, ITCH, IYD, KANSL1, KAT6B, KCNJ1, KCNJ10, KCNJ2, KDM5C, KDM6A, KIF1A, KIF22, KIF7, KLF1, MLL, MLL2, KRAS, L1CAM, LAMTOR2, LARGE, LEPRE1, LHX3, LHX4, LIFR, LMNA, LMX1B, LRBA, LRP2, LRP5, LTBP2, LTBP3, LTBP4, MAN2B1, MAP2K1, MAP2K2, MASP1, MATN3, MBTPS2, MCPH1, MECP2, MED12, MGAT2, MGP, MIR17HG, MKS1, MLYCD, MMP13, MMP2, MMP9, MPO, MPV17, MVK, MYCN, MYH3, MYH8, NAA10, NBAS, NBN, NDE1, NDN, NEK1, NF1, NIN, NIPBL, NKX2-1, NKX2-5, NOG, NOTCH2, NPR2, NSD1, NSDHL, NSUN2, OCRL, OFD1, ORC1, ORC4, ORC6, OTX2, PAPSS2, PAX8, PCCB, PCNT, PDE4D, PEX7, PFKM, PGAM2, PHEX, PHF6, PHGDH, PHKA1, PHKA2, PHKB, PIGO, PIK3R2, PITX2, PLEC, PLOD2, PLOD3, POC1A, POR, PORCN, POU1F1, PPIB, PQBP1, PRKAR1A, PROP1, PTH1R, PTHLH, PTPN11, PYCR1, RAB23, RAB3GAP1, RAB3GAP2, RAB40AL, RAD21, RAD51C, RAF1, RAPSN, RARA, RBBP8, RBM8A, RECQL4, RIN2, RIPK4, RMRP, RNU4ATAC, ROR2, RPS19, RPS6KA3, RTTN, SBDS, SDHA, SEC23A, SECISBP2, SEMA3E, SEPN1, SEPT9, SERPINF1, SERPINH1, SF3B4, SH3BP2, SHH, SHROOM4, SIL1, SLC12A1, SLC16A2, SLC17A5, SLC19A2, SLC26A2, SLC26A4, SLC29A3, SLC34A1, SLC34A3, SLC35C1, SLC35D1, SLC37A4, SLC39A13, SLC39A4, SLC5A5, SLC6A19, SLC6A8, SLC9A6, SLX4, SMAD4, SMARCA2, SMARCAL1, SMC1A, SMC3, SMOC1, SMPD1, SMS, SNRPN, SOS1, SOST, SOX2, SOX3, SOX9, SP7, SPG20, SRCAP, STRA6, SUMF1, TAZ, TBCE, TBX1, TBX15, TBX3, TCTN3, TERT, TFAP2A, TG, THRA, THRB, TINF2, TMCO1, TMEM165, TMEM237, TNFRSF11B, TNNI2, TNNT3, TP63, TPM2, TPO, TRAPPC2, TRH, TRHR, TRIM37, TRIP11, TRPS1, TRPV4, TSHB, TSHR, TWIST1, UBR1, UROC1, UROS, USB1, VDR, VLDLR, VPS13B, WDR19, WDR35, WFS1, WISP3, WNT5A, WNT7A, WRN, ZBTB16, ZEB2, ZMPSTE24 and ZNF592.
3. the test kit of claim 1 or 2, it is characterised in that, described probe can also specific recognition its can the intron region of each 30bp of upstream and downstream of exon region of each gene of specific recognition.
4. the test kit of claim 1 or 2 is in the purposes detected in short and small genes involved.
5. the method for the short and small genes involved of detection, it is characterised in that, comprising:
(1) obtaining the nucleic acid of person under inspection, described nucleic acid is genomic nucleic acids and/or free nucleic acid fragment;
(2) catch described nucleic acid, obtain short and small genes involved region;
(3) described short and small genes involved region is carried out sequencing, obtain sequence information;
(4) described short and small genes involved is detected based on described sequence information;
Wherein, (2) utilize the arbitrary test kit of claim 1-3 to carry out.
6. the method for claim 5, it is characterised in that, (4) comprise based on described sequence information detect simultaneously described short and small genes involved SNP and micro-deleted micro-repeat variation.
7. the method for claim 6, it is characterised in that, the SNP and micro-deleted micro-repetition that detect described short and small genes involved make a variation simultaneously, comprising:
Described sequence information and reference sequences are carried out the first comparison, obtains the first comparison result;
A part for described first comparison result and described reference sequences is carried out the 2nd comparison, obtains the 2nd comparison result;
Based on described first comparison result and described 2nd comparison result, the SNP and micro-deleted micro-repetition that detect described short and small genes involved make a variation simultaneously.
8. the method for claim 7, it is characterised in that, described first comparison is overall comparison, and described 2nd comparison is local comparison.
9. the method for claim 7, it is characterised in that, described reference sequences is hg19.
10. the method for claim 7, it is characterized in that, a part for described reference sequences comprises the known micro-deleted micro-repetition site of each in the described reference sequences corresponding with described short and small genes involved region, and the reference sequences of each known micro-deleted micro-repetition site each 1000bp of upstream and downstream described.
The method of 11. claims 7, it is characterised in that, utilize GATKUnifiedGenotyper software detect simultaneously described short and small genes involved SNP and micro-deleted micro-repeat variation.
The device of 12. 1 kinds of short and small genes involveds of detection, it is characterised in that, comprising:
A. nucleic acid acquiring unit, for obtaining the nucleic acid of person under inspection, described nucleic acid is genomic nucleic acids and/or free nucleic acid fragment;
B. capturing unit, is connected with A unit, for catching from the nucleic acid in A unit, to obtain short and small genes involved region;
C. sequencing unit, is connected with unit B, for the short and small genes involved region from unit B is carried out sequencing, to obtain sequence information;
D. detecting unit, is connected with C cell, for based on the sequence information described short and small genes involved of detection from C cell; Wherein, catching in unit B utilizes the arbitrary test kit of claim 1-3 to carry out.
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| CN110438222A (en) * | 2018-05-04 | 2019-11-12 | 中国科学院上海生命科学研究院 | A kind of early diagnosis detection kit for aggressive lymphomas |
| CN110724710A (en) * | 2019-10-28 | 2020-01-24 | 扬州大学 | Vector for controlling pig PFKM expression and application thereof |
| CN115873937A (en) * | 2022-08-15 | 2023-03-31 | 苏州市立医院 | Biomarker for predicting occurrence of repeated planting failure and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2005089436A2 (en) * | 2004-03-16 | 2005-09-29 | The Regents Of The University Of California | Genetic networks regulated by attenuated gh/igf1 signaling and caloric restriction |
| CN101058832A (en) * | 2000-05-12 | 2007-10-24 | 加的夫大学学院咨询有限公司 | Method for detecting growth hormone variations in humans, the variations and their uses |
| CN102329876A (en) * | 2011-10-14 | 2012-01-25 | 深圳华大基因科技有限公司 | Method for measuring nucleotide sequence of disease associated nucleic acid molecules in sample to be detected |
-
2014
- 2014-11-13 CN CN201410639462.8A patent/CN105648043A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101058832A (en) * | 2000-05-12 | 2007-10-24 | 加的夫大学学院咨询有限公司 | Method for detecting growth hormone variations in humans, the variations and their uses |
| WO2005089436A2 (en) * | 2004-03-16 | 2005-09-29 | The Regents Of The University Of California | Genetic networks regulated by attenuated gh/igf1 signaling and caloric restriction |
| CN102329876A (en) * | 2011-10-14 | 2012-01-25 | 深圳华大基因科技有限公司 | Method for measuring nucleotide sequence of disease associated nucleic acid molecules in sample to be detected |
Non-Patent Citations (3)
| Title |
|---|
| MICHAEL H. GUO,ET AL: "Whole Exome Sequencing to Identify Genetic Causes of Short Stature", 《HORM RES PAEDIATR》 * |
| STEVEN D. CHERNAUSEK: "Whole Exome Sequencing in Short Stature: Finding Needles in the Haystack", 《HORM RES PAEDIATR》 * |
| 于萍: "68例矮小症患者的基因变异检测和病因研究", 《中国优秀硕士学位论文全文数据库》 * |
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| CN106834509B (en) * | 2017-03-20 | 2020-10-16 | 济宁医学院附属医院 | Application of primer and reagent in preparation of reagent group for screening growth hormone deficiency |
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| CN110272994B (en) * | 2019-07-15 | 2021-01-26 | 中国医学科学院北京协和医院 | Gene mutation diagnosis of CVM and application thereof |
| CN110724710A (en) * | 2019-10-28 | 2020-01-24 | 扬州大学 | Vector for controlling pig PFKM expression and application thereof |
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