19 results about "Reference gene" patented technology

The invention relates to the technical field of molecular biology and environmental biology, and discloses a PCR (Polymerase Chain Reaction) chip and a kit for detecting the expression quantity of functional genes related to microbial degradation of chlorinated hydrocarbon and application of the PCR chip and the kit. The PCR chip comprises a primer pair and a primer pair, the primer pair is used for specifically amplifying dehalogenation functional genes bvcA, cfrA, mbrA, pceA, pteA, tceA and vcrA respectively, the primer pair is used for specifically amplifying reference genes, and nucleotide sequences of the primer pair for specifically amplifying the dehalogenation functional genes are shown as SEQ ID NO. 1 to SEQ ID NO. 14. The PCR chip provided by the invention can specifically amplify seven functional genes related to the degradation of the chlorinated hydrocarbon microorganisms, is good in specificity, high in accuracy and good in repeatability, and can accurately detect the expression condition of the seven dehalogenation functional genes related to the degradation of the chlorinated hydrocarbon microorganisms through a qPCR technology, so that the existence condition of chlorinated hydrocarbon degrading bacteria can be reflected; the method has reliability in the aspects of chlorohydrocarbon polluted site biodegradability evaluation, degradation process research, repair effect verification and the like.

The invention provides an NOD / shiLt mouse genotype fluorescent quantitative identification method and application, and relates to the field of experimental animal genetic quality control. Comprising the following steps: acquiring a to-be-detected mouse genetic identification sample, and performing DNA extraction treatment on the mouse genetic identification sample to form a genome DNA template; according to the fluorescent quantitative identification method for the genotype of the NOD / shiLt mouse and the application of the fluorescent quantitative identification method, a real-time fluorescent quantitative polymerase chain reaction system containing a specific primer pair, an insertable fluorescent dye premixed solution and a standard substance series is constructed, so that accurate quantitative detection on a specific deletion sequence of a No.3 chromosome of the NOD / shiLt mouse and a reference gene GAPDH is realized. According to the present invention, the linear relationship between the Ct value and the template copy number is established through the standard substance series, and the quality determination of the amplification curve, the melting curve and the reference gene is combined, such that the NOD / shiLt mouse genotype fluorescence quantitative identification method can stably and rapidly identify whether the mouse genomic DNA contains the NOD / shiLt mouse genotype.

The invention provides a reference gene combination for real-time fluorescent quantitative PCR (Polymerase Chain Reaction) of indocalamus as well as a primer pair combination and application of the reference gene combination, and belongs to the technical field of plant molecular biology. In combination with indocalamus transcriptome data analysis and cross validation of four algorithms, reference genes, MD10B, PP2A, eIF1A, Ite23725, eIF4A, 60S and UBP1, suitable for indocalamus are systematically screened out, optimal reference gene combinations under different experimental conditions (drought, salt, waterlogging stress and different tissues) are refined, and the method is scientific and rigorous and has good application prospects. And a precise tool is provided for gene expression analysis of the indocalamus in different research directions. The screened reference genes show extremely high expression stability in specific stress and tissues after being combined, and the expression stability is far better than that of traditional housekeeping genes such as Actin and UBI. The specific primer has high amplification efficiency, and ensures the accuracy of qRT-PCR results.

The application belongs to the technical field of molecular diagnosis, and particularly relates to a digital PCR kit and method for accurately detecting the copy number of a CAR gene of a CAR-T cell and a lentivirus vector. The application first realizes the absolute quantification of CAR, Rev and an internal reference gene RNase P in a single tube simultaneously, and has strong universality. Based on the principle of digital PCR, the absolute copy number is directly obtained without a standard curve, and the result is accurate and highly repeatable. The detection lower limit reaches 0.01%, and extremely low abundance CAR-T residues or microresidual diseases can be detected. Primers and probes are designed for the conservative regions of CD3zeta, Rev and RNase P, and no cross reaction is verified, so that false positives are effectively avoided. The sample only needs gDNA, and there is no requirement for cell activity, and the sample is compatible with freezing and transportation. The kit is premixed and optimized, the process is automated, and 4-5 hours are needed. The kit is suitable for the whole chain of CAR-T drug research and development, production process quality control, clinical patient monitoring and safety evaluation.

The invention discloses a composition for detecting spinal muscular atrophy typing based on nucleic acid mass spectrometry and application of the composition. The invention relates to the field of gene detection, and provides a composition for detecting spinal muscular atrophy typing based on nucleic acid mass spectrometry, which comprises a target gene primer pair, a reference gene primer pair and an IIS type restriction enzyme. The composition can be used for detecting spinal muscular atrophy typing based on nucleic acid mass spectrometry, and the traditional technical method of nucleic acid mass spectrometry is improved, so that nucleic acid mass spectrometry detection does not need the steps of SAP treatment and single base extension reaction, the reaction time is shortened, the result is accurate and reliable, the steps are simple, and the cost is greatly reduced. The invention is of great significance to preparation of screening products suitable for large-scale screening of spinal muscular atrophy.

The invention relates to the technical field of kits, and discloses a peanut allergen component fluorescence PCR (Polymerase Chain Reaction) detection equivalent kit, which comprises (1) 2 * Probe qPCR Mix; (2) a florescence source probe primer mixture; (3) a 18S internal reference probe primer mixture; (4) ultrapure water; (5) positive control; (6) negative control; according to the invention, a dual detection mechanism of a florescence source specific gene and an eukaryote general reference gene is combined, and a specially designed chimeric recombinant plasmid is adopted as a multifunctional positive control, so that when a complex food matrix is treated, a forced standard reaching mechanism of a reference gene channel can timely identify a sample treatment abnormal condition; and wrong interpretation caused by improper operation is avoided. Meanwhile, the activity of core components of the kit can be verified through the two-channel synchronous response characteristic of positive control, and the credibility of a detection result and the scientific preciseness of interpretation are improved.

This invention discloses a one-step RT-PCR method for detecting five viruses in *Calamus sylvestris*, belonging to the field of plant virus molecular detection technology. Specifically, it provides a primer set for the simultaneous detection of multiple viruses in *Calamus sylvestris*, including five pairs of virus-specific primers for the specific detection of PVBV, CaYMAV, CaYMV, CaYSV, and SCMV, and one pair of *Calamus sylvestris* internal reference gene primers, the sequences of which are shown in SEQ ID NO:1-12. RNA is extracted from *Calamus sylvestris* samples, a one-step RT-PCR reaction system is prepared, amplification and electrophoresis are performed, the detection validity is verified by the internal reference gene band, and the type of virus infection is determined by the virus-specific bands. This invention can achieve simultaneous detection of five viruses and has the advantages of simple operation, short detection cycle, high sensitivity, and strong specificity, making it suitable for rapid screening of large batches of *Calamus sylvestris* samples.

The invention discloses a digital PCR detection system, kit and method for detecting single nucleotide polymorphism and copy number variation of a CYP2D6 gene and application, and belongs to the technical field of molecular biology. The method comprises the following steps: designing three pairs of CYP2D6 and reference gene RPP30 specific primers, matching with four locked nucleic acid modified TaqMan fluorescent probes, constructing a single-tube multiple digital PCR detection system, and synchronously and accurately detecting three single nucleotide polymorphisms of CYP2D6 * 10, CYP2D6 * 36 and CYP2D6 * 41 and CYP2D6 * 5 copy number variation; wherein the RPP30 reference gene can synchronously realize sample amplification quality control and target gene absolute quantitative calibration. The detection system depends on the advantages of high specificity of the LNA probe and high sensitivity of digital PCR, the detection sensitivity reaches 0.02 ng / mu L, the detection period is short, and the result is stable and reliable.

The invention relates to a real-time quantitative polymerase chain reaction (qPCR) method and a kit, which are used for simultaneously determining classification of specific gene variation and gene copy number. More specifically, the present invention relates to an efficient qPCR method and kit for simultaneously determining the variation and copy number of a target gene by performing real-time quantitative PCR on a reference gene and a target gene with known copy number in the same reaction vessel.

The invention discloses a method for detecting activity of human induced pluripotent stem cell derived myocardial cell telomerase, a reference gene, a primer and probe combination and application, and belongs to the technical field of stem cell biology and molecular detection. According to the method, stably expressed 18s rRNA is taken as a reference gene, total RNA at different differentiation stages of human induced pluripotent stem cell derived myocardial cells is extracted and is subjected to reverse transcription into cDNA, then a specific primer probe combination is adopted, the gene expression quantity of telomerase reverse transcriptase and 18s rRNA is detected through Taqman fluorescent quantitative PCR, and the activity of telomerase is analyzed and evaluated in a normalized manner in combination with a 2-delta delta CT method. The method solves the problem of detection deviation caused by instability of traditional reference genes, and is simple and convenient to operate, good in repeatability and reliable in result. The method can be used for evaluating the differentiation efficiency of the human induced pluripotent stem cell derived myocardial cells, the maturity of the myocardial cells, drug screening and heart disease model research, and has important application value.

The application particularly relates to a primer probe, a kit and application for detecting the GJB2 gene copy number of a non-syndromic hearing loss patient by using microdroplet digital PCR. The primer probe combination provided by the application comprises a target gene detection primer pair, a target gene detection probe, an upstream primer of a reference gene, a downstream primer of the reference gene and a reference gene probe, and the sequence information is shown as SEQ ID NO. 6-11; and the application further provides a kit for detecting the GJB2 gene copy number of a non-syndromic hearing loss patient. The primer probe combination and the kit provided by the application are applied to the detection of the GJB2 gene copy number variation of a non-syndromic hearing loss patient, and the accuracy of the detection of genetic hearing loss is significantly improved; according to the determination result, it is determined whether the GJB2 gene expression is abnormal or not, the disease prognosis is evaluated, potential therapeutic drugs are screened, genetic counseling services are provided or individualized medical schemes are formulated, and the application has a wide application prospect.

The present application relates to the field of C12Q1 / 6851, and more particularly, the present application relates to a primer probe combination, a kit and a method for detecting human ALK and ROS1 gene fusion. The primer probe combination comprises at least a primer probe combination A which specifically detects at least one EML4-ALK fusion gene; a primer probe combination B which specifically detects at least one ROS1 fusion gene; and a primer probe combination C which specifically detects an internal reference gene. The primer probe combination and the detection method of the present application have high specificity and sensitivity, improve the signal-to-noise ratio, have good resolution and accuracy, and have excellent linear relationship, so that the result can be easily judged.

The invention provides an automatic single cell annotation method and system, and relates to the technical field of automatic cell type annotation. According to the method, a reference gene set is constructed based on marker genes of a reference map, a weight rule is set, and a probability score and confidence of a cell type are calculated in combination with a semi-supervised, unsupervised or predefined module mode. Through multi-algorithm integration and reannotation rechecking, annotation precision and consistency are optimized, final labels and related evidences are output, and data tracing and reanalysis are supported.

The invention relates to the technical field of molecular biology and environment, and discloses a primer group, a kit and application thereof, and a method for detecting biotoxicity of an environmental sample, the primer group comprises a primer pair for specific amplification of an hsp70.3 gene and an mt2 gene, and a primer pair for specific amplification of a reference gene; the primer pair for specifically amplifying the hsp70.3 gene comprises an upstream primer with a nucleotide sequence as shown in SEQ ID NO.3 and a downstream primer with a nucleotide sequence as shown in SEQ ID NO.4; the primer pair for specifically amplifying the mt2 gene comprises an upstream primer with a nucleotide sequence as shown in SEQ ID NO.5 and a downstream primer with a nucleotide sequence as shown in SEQ ID NO.6. The primer group disclosed by the invention can be used for monitoring the content change of hsp70.3 and mt2 in a living body, so that the biotoxicity of a to-be-detected environmental sample can be analyzed, the early warning of heavy metal pollutants is realized, particularly the existence of the heavy metal pollutants in the to-be-detected environmental sample can be prompted, and the heavy metal detection can be further combined for analysis in the later period.

The application relates to the fields of molecular biology and environmental biotechnology, and discloses a PCR chip, a kit and application thereof. The PCR chip comprises a chip plate, primer pairs for specifically amplifying bvcA genes, cfrA genes, mbrA genes, pceA genes, pteA genes, tceA genes, vcrA genes and internal reference genes are loaded on the chip plate, and a gradient dilution template of an internal reference gene for drawing a standard curve is arranged on the chip plate; the gradient dilution template of the internal reference gene is obtained by gradient dilution of internal reference gene template DNA and addition of internal reference gene primers. The PCR chip can detect absolute quantification of chlorinated hydrocarbon microbial degradation related reduction dehalogen functional genes through qPCR technology, so that the PCR chip can be used for accurately detecting the copy number of the chlorinated hydrocarbon microbial degradation related reduction dehalogen functional genes, thereby the feasibility of chlorinated hydrocarbon pollution bioremediation can be explored, the chlorinated hydrocarbon pollution bioremediation process can be evaluated, and the chlorinated hydrocarbon pollution bioremediation method can be further assisted and guided.

The invention discloses a methylation marker combination for early screening of endometrial cancer, and belongs to the cross technical field of molecular diagnosis and bioinformatics. Three methylation markers CDO1, NKX2-6 and SORCS3 are screened out through bioinformatics analysis, NGS targeted sequencing and qPCR verification, specific primers SEQ ID NO.1-2, 4-5 and 7-8 and probes SEQ ID NO.3, 6 and 9 of the methylation markers are designed, and a fluorescent quantitative PCR detection system is constructed in combination with a reference gene BACT. According to the method, the methylation level of a three-gene promoter region in a sample is detected, the cutoff value is 0.36, the sensitivity of endometrial cancer diagnosis of cast-off cells is 90.37%, the specificity is 89.60%, AUC is equal to 0.9682, and benign and cancerous samples can be effectively distinguished. According to the invention, noninvasive / minimally invasive early screening of endometrial cancer is realized, diagnosis accuracy and safety are improved, and NKX2-6 and SORCS3 are newly discovered markers and have important scientific value and clinical transformation potential.

The invention relates to the technical field of molecular biology, in particular to an SNP (Single Nucleotide Polymorphism) molecular marker for resisting fusarium verticillium ear rot of corn, application and a detection method. According to the SNP molecular marker, KA002834 is located at the position of 156327595bp of a No.3 chromosome of a reference gene sequence AGPv5 of corn, the disease-resistant allele site is T, and the susceptible allele site is A; kA002839 is located at the position of No.3 chromosome 157801329bp of a reference gene sequence AGPv5 of corn, the disease-resistant allele site is C, and the susceptible allele site is A. The SNP molecular marker is closely linked with the maize fusarium verticillium ear rot resistant gene, the recombination rate is low, the marker stability is high, and the anti-disease gene can be tracked; the disease-resistant genotype can be identified through DNA detection in the corn seedling stage, so that the field production cost is reduced, the interference of the environment on phenotype identification is avoided, and the breeding period is effectively shortened.

The application discloses a real-time fluorescent quantitative PCR internal reference gene of Onychostoma rhabarbarium and a screening method and application thereof. The application is based on Onychostoma rhabarbarium genome assembly and gene annotation work, and 18 candidate internal reference genes are screened according to transcriptome data of 10 different tissues. The expression of the 18 candidate internal reference genes in different tissues of the Onychostoma rhabarbarium and the stability of the expression in different tissues are detected by a qPCR method, and the result shows that, tubb4b , eif2s2 , eif3a or rps27l the expression is most stable in various tissues, and the combination of the internal reference genes is recommended as the internal reference gene group of the qPCR experiment of the Onychostoma rhabarbarium. The result of the application lays a foundation for functional gene research of the Onychostoma rhabarbarium, especially expression mode research of the functional gene in various tissues, and provides a molecular biology basis for resource protection and artificial breeding of the Onychostoma rhabarbarium.

This invention discloses the application of AGR2, TFF1, and EGFR genes as a combination of biomarkers in the preparation of gastric cancer diagnostic reagents or kits. The reagents or kits are based on... AGR2, TFF1, EGFR Genes and internal reference genes ACTB Design specific PCR primers and TaqMan probes. Detected in cell-free DNA in plasma. AGR2 Genes, aiding in the prediction of moderately to poorly differentiated gastric cancer; detected in cDNA samples synthesized from plasma exosomal mRNA via reverse transcription. AGR2 Genes, aiding in the prediction of metastasis in moderately to poorly differentiated gastric cancer; in cDNA samples synthesized from mRNA reverse transcribed from ascites exfoliated cells and gastric muscularis propria tissue, simultaneously satisfying... AGR2 Gene △ Ct （AGR2‑ACTB） <6 and AGR2 TFF1 Gene △ Ct （TFF1‑ACTB） <6, assisting in the rapid diagnosis of moderately or poorly differentiated gastric cancer. Compared with existing protein level detection methods, this invention has higher detection sensitivity and timeliness.