82 results about "Reference gene" patented technology

The invention provides a method and a device for designing a protein coding codon sequence based on a large multi-modal model, which comprehensively utilize information such as an amino acid sequence, space folding and mRNA quality of protein to realize high-quality, controllable and evaluable codon sequence design. According to the method, the dependence of a traditional codon optimization strategy on a reference gene template is broken through, and a codon optimization problem is converted into a codon design task based on protein information. Through comprehensive consideration of information such as space folding of protein, context of amino acid sequences, mRNA quality and the like, the model can generate a codon sequence which is more in line with codon use preference of specific species, reasonable in structure and efficient in expression, so that a new thought and technical support are provided for safe, stable, efficient and controllable protein expression.

The invention provides a primer probe combination and a kit for detecting the copy number of a 22q11 region based on digital PCR and a use method of the primer probe combination. Specific probes of a target gene TBX1, a target gene CRKL, a target gene SMARCB1 and a reference gene GAPDH are integrated in different fluorescence channels, and the copy number of the target gene is detected according to the ratio of the copy number of the reference gene to the copy number of the three target genes. According to the invention, amplification products of different sites in different fluorescence channels are integrated in the same system in digital PCR reaction, so that the detection flux is greatly improved, and low-cost and high-flexibility accurate quantification of 22q11. 2 copy number variation is realized through a single-channel multi-target fluorescence layering strategy; and the dependence on expensive equipment, complex operation and professional analysis in the prior art is overcome.

The invention discloses an endogenous reference gene, a primer probe and a kit for quantitatively detecting HBV (Hepatitis B Virus) nucleic acid. The nucleotide sequence of the endogenous reference gene is shown as SEQ ID NO: 1 or SEQ ID NO: 2. Two endogenous reference gene sequences capable of being used for quantitatively detecting HBV nucleic acid and several groups of primer probes for endogenous reference gene detection and HBV nucleic acid gene detection are obtained through screening, quantitative detection of HBV nucleic acid can be rapidly and accurately achieved through the primer probes, the primer probes can be used for detecting HBV genotype A-I, and the detection range is wider.

The invention discloses a pig actinobacillus pleuropneumoniae constant-temperature detection primer group capable of avoiding a false negative detection result and application of the pig actinobacillus pleuropneumoniae constant-temperature detection primer group, the pig actinobacillus pleuropneumoniae constant-temperature detection primer group comprises an xerD detection primer group and an internal standard primer group, and the detection primer group is an LAMP primer group taking a xerD gene of the pig actinobacillus pleuropneumoniae as a target gene, comprising an outer primer with a nucleotide sequence as shown in SEQ ID NO.1-2, an inner primer with a nucleotide sequence as shown in SEQ ID NO.3-4 and a loop primer with a nucleotide sequence as shown in SEQ ID NO.5-6. The internal standard primer group is an LAMP (Loop-Mediated Isothermal Amplification) primer taking an actinobacillus pleuropneumoniae apxIVA gene as an internal standard gene, and comprises outer primers with nucleotide sequences as shown in SEQ ID NO.7-8, inner primers with nucleotide sequences as shown in SEQ ID NO.9-10 and loop primers with nucleotide sequences as shown in SEQ ID NO.11-12. The method has the advantages that the detection time is short, the operation is easy, the endogenous reference gene is added on the basis of the isothermal amplification technology, the false negative is effectively identified, the detection accuracy of the actinobacillus pleuropneumoniae is facilitated, and the popularization and application of the constant-temperature detection method in detection are facilitated.

The invention provides a primer probe composition and a kit for post-operation MRD (minimal residual focus) detection and dynamic monitoring of pancreatic ductal adenocarcinoma (PDAC). The primer probe composition disclosed by the invention is used for amplifying a CpG site enrichment region of a gene mir129-2 promoter region. By using the method provided by the invention, the specific CpG site enrichment region can be specifically amplified in a high methylation state, and cannot be amplified in a low or no methylation state. The kit provided by the invention comprises a methylation primer and a probe of the gene mir129-2, and further comprises a primer and a probe of a reference gene ACTB. The kit can be used for detecting the methylation state of the gene mir129-2 starter region, and the methylation state of the gene mir129-2 starter region can be used as a marker for post-operation MRD detection and dynamic monitoring of pancreatic ductal adenocarcinoma (PDAC), and is used for evaluating the recurrence risk of a post-operation patient of the pancreatic ductal adenocarcinoma (PDAC).

The invention discloses a primer probe combination and a kit for on-site detection of a transgenic maize transformant, and relates to the technical field of transgenic detection. The invention provides a primer probe combination for on-site detection of a transgenic maize transformant. The primer probe combination comprises a primer pair and a probe for detecting a transgenic maize strain MON810, the primer pair and the probe are used for detecting the transgenic maize strain TC1507; the primer pair and the probe are used for detecting the transgenic maize strain Bt11; the invention discloses a primer pair and a probe for detecting corn internal reference zSSIIb. The method for detecting the transgenic maize by adopting the primer probe combination and the kit disclosed by the invention integrates sample pretreatment, nucleic acid extraction, purification, amplification and detection, has better sensitivity and accuracy, is simple, convenient, rapid and time-saving to operate, and plays an important role in on-site rapid detection of transgenic maize research and safety management.

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 discloses a primer probe and a kit for detecting H3F3A gene mutation and its application, wherein the H3F3A gene mutation type is K27M, G34R and G34V mutation. The kit comprises a set of primer probe groups for detecting H3F3A gene mutation and a universal reference gene primer probe group, wherein the primer probe group for detecting H3F3A gene mutation comprises specific upstream primers for K27M, G34R and G34V mutations, a universal downstream primer and a universal fluorescent probe respectively. The kit provided by the invention can detect common mutation types H3F3A K27M, H3F3A G34R and H3F3A G34V in the H3F3A gene, has the advantages of high sensitivity, convenient and fast operation and low cost, and has broad clinical application prospects in diagnosing diseases such as diffuse midline glioma and diffuse cerebral hemispheric glioma.

The invention discloses a detection method for detecting enteropathogenic bacteria, and relates to the field of molecular biology, specific genes are searched in genomes of salmonella, shigella, vibrio parahaemolyticus, campylobacter jejuni, diarrheagenic escherichia coli, vibrio cholerae and staphylococcus aureus, a beta-actin gene is adopted as a reference gene, and the enteropathogenic bacteria are detected. Primer and probe sequences are designed, a probe group consisting of the primer and probe sequences is used for carrying out real-time fluorescent PCR amplification reaction on nucleic acid in a detected sample, then melting curve analysis is carried out, and whether the pathogenic bacteria exist in the nucleic acid sequence to be detected or not can be judged according to a Tm value. The detection method provided by the invention has the advantages of rapidness, high throughput, accuracy and high sensitivity.

The invention relates to an internal reference gene of solanum muricatum under verticillium wilt stress, a primer and application, and belongs to the technical field of solanum muricatum molecular biology. The nucleotide sequence of the reference gene Ss18SRNA is as shown in SEQ ID NO. 1. The nucleotide sequences of RT-qPCR (real-time quantitative polymerase chain reaction) amplification primers of the reference gene Ss18SRNA are as follows: a forward primer is as shown in SEQ ID NO.8; and the reverse primer is as shown in SEQ ID NO.9. The reference gene is stably and consistently expressed under different time conditions after the solanum melongena is inoculated with verticillium wilt pathogenic bacteria, can be used for analyzing and evaluating the expression condition of related functional genes under the verticillium wilt stress condition, and provides a reference gene for quantitative analysis of gene expression of solanum melongena under the verticillium wilt stress condition.

The invention discloses a reference gene for acer ginnala gene expression analysis as well as a primer and application thereof, and belongs to the technical field of plant molecular biology. The reference gene is AgTUA and / or AgPP2A and / or AgACT2, the nucleotide sequence of the AgTUA gene is as shown in SEQ ID NO.4, the nucleotide sequence of the AgPP2A gene is as shown in SEQ ID NO.6, and the nucleotide sequence of the AgACT2 gene is as shown in SEQ ID NO.2. The invention further discloses a kit for detecting the content of the AgPP2A gene and the AgACT2. The invention also provides amplification primers for detecting the reference gene, and the primer sequences of the amplification primers are respectively shown as SEQ ID NO.16 and SEQ ID NO.17, SEQ ID NO.20 and SEQ ID NO.21, and SEQ ID NO.12 and SEQ ID NO.13. The invention also provides a kit for detecting the reference gene. The reference gene is suitable for real-time fluorescent quantitative expression analysis of acer ginnala flavonoid synthesis related genes or other functional genes, the current situation that acer ginnala has no reference gene is filled, and the accuracy and reliability of gene expression analysis data are improved; powerful support is provided for acer ginnala functional gene expression analysis and regulation mechanism research.

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 belongs to the technical field of virus detection, and particularly relates to a double-channel real-time fluorescent quantitative PCR (Polymerase Chain Reaction) detection primer, a probe, a detection kit, a detection method and application thereof. The detection sensitivity is high, and the lowest detection limit can reach 10 copies / mL; the detection time period is short, nucleic acid detection can be completed within 45 minutes, and the kit is suitable for clinical and bedside rapid detection and diagnosis; the detection specificity is high, and cross amplification with other 80 pathogens is avoided; the detection accuracy is 100%; virus quantitative analysis can be carried out, and the quantitative linear range is good; the detection repeatability is good, and the precision is high; uDG enzyme is added into a detection system, so that pollution can be prevented; a reference gene is also added into the detection system, so that the quality of the whole extraction and amplification process of the sample can be monitored according to the detection result of the reference gene, and the existence of the RNA of the Enduma virus in samples such as whole blood, serum, plasma, cerebrospinal fluid, urine, alveolar lavage fluid, throat swab and excrement can be detected.

The invention discloses a reference gene suitable for gene expression analysis of bombyx mori in a full development stage, a screening method, primers and application, and belongs to the technical field of molecular biology. A reference gene CYPA is screened out through a transcriptomics technology, the expression quantity of the reference gene CYPA in all development stages of bombyx mori is larger than 190, the expression quantity is stable and has no significant difference, primer sequences, shown as SEQ ID No: 2 and SEQ ID No: 3, applied to qPCR experiments are designed according to a CYPA gene sequence, and the CYPA primer sequences have the advantage of being high in specificity and can be applied to detection of the full development stages of bombyx mori; according to the technical scheme, the defect that reference genes are selected according to experience in different development stages of bombyx mori is overcome, the stable reference genes are provided for gene expression analysis in the full development stage of bombyx mori, and the accuracy and credibility of qPCR results of related genes in the full development stage of bombyx mori are improved.

The invention relates to a method for inferring a silkworm chrysalis stage development process based on trimethyl lysine hydroxylase. The method comprises the following steps: (1) constructing a correlation model of relative transcription expression quantity of a trimethyl lysine hydroxylase gene and a reference gene of a silkworm chrysalis sample of a variety to be detected and pupal stage development time; and (2) determining the relative transcription expression quantity of the trimethyl lysine hydroxylase and the reference gene in the silkworm chrysalis to be detected, and calculating the pupal stage development time according to the correlation model obtained in the step (1). According to the method, the development process of one or more silkworm chrysalis can be accurately measured, errors caused by traditional visual inspection can be reduced, a new detection method is provided for deducing the pupal development process of the silkworms, and an important basis is provided for judging moth formation time in silkworm seed production work.

The invention relates to the technical field of nucleic acid detection, in particular to an endogenous reference gene, a probe primer and a kit for quantitatively detecting HCMV nucleic acid, the endogenous reference gene comprises a core sequence and flanking sequences located at the two ends of the core sequence, and the nucleotide sequence of the core sequence is shown as SEQ ID NO.1. According to the endogenous reference gene sequence, the detection primer and the detection probe of the endogenous reference gene and the detection primer and the detection probe for specifically recognizing the HCMV nucleic acid, rapid and accurate quantitative detection of the HCMV nucleic acid can be realized, and the accuracy and the reliability of a detection result are improved.

The present invention provides a method for detecting the insertion copy number of foreign genes in Setaria viridis or foxtail millet by fluorescence quantitative PCR or digital PCR, belonging to the field of biotechnology. The method uses RA1 gene as an internal reference gene and Hyg screening marker gene as a foreign gene, and uses primer pairs RA1-F / RA1-R and primer pairs Hyg-F / Hyg-R to detect the genomic DNA of transgenic Setaria viridis or transgenic foxtail millet, and determine the insertion copy number of foreign genes in transgenic Setaria viridis or transgenic foxtail millet. The present invention provides a method for detecting the insertion copy number of foreign genes in Setaria viridis or foxtail millet by fluorescence quantitative PCR or digital PCR, which has the characteristics of simplicity, rapidity, low sample consumption, high efficiency, high throughput, etc., can improve the efficiency of screening transgenic lines of Setaria viridis or foxtail millet with low-copy insertions, and provides a new option for the detection method of the insertion copy number of foreign genes in the research of transgenic Setaria viridis or transgenic foxtail millet.

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 present invention provides a kit for realizing human NK lymphocyte counting based on gene methylation detection, and the kit is a kit for detecting methylation of killer cell immunoglobulin-like receptor 2DL4 (KIR2DL4) gene in a sample. The present invention proposes for the first time that the unmethylated gene region of KIR2DL4 gene can be used as a biomarker for NK cell identification. The present invention quantifies the copy number of KIR2DL4 target gene and internal reference gene by PCR technology, and can conveniently and quickly perform relative quantitative analysis on NK cells in a small amount of blood or dried blood film. The target detected by the present invention is the methylation level of leukocyte DNA in the sample, and can be applied to various types of samples, including fresh or frozen blood, isolated PBMC cells, dried blood spots, etc., and the requirements for samples are greatly reduced.

The present invention discloses an internal reference gene of a fruit of Phyllanthus emblica at different developmental stages, a detection primer and an application thereof, wherein the internal reference gene is PeSTRG.26181.1.p1, and its nucleotide sequence is shown in SEQ ID NO.13. The present invention preliminarily screens out 13 candidate internal reference genes through transcriptome sequencing data, and then screens out the internal reference gene PeSTRG.26181.1.p1 most stably expressed in Phyllanthus emblica fruit from the 13 candidate internal reference genes based on five internal reference gene stability evaluation software. The present invention also provides real-time fluorescence quantitative PCR detection primers for the internal reference gene, and the primer sequences are shown in SEQ ID NOs.38 and 39. The internal reference gene and the detection primer can be used in the real-time fluorescence quantitative expression analysis of Phyllanthus emblica functional genes, which can significantly improve the specificity, stability and accuracy of functional gene expression data, and provide strong support for the expression analysis of Phyllanthus emblica functional genes.

The present invention discloses a molecular marker combination for detecting adulterated Bos genus milk in buffalo milk and its application, including molecular marker 1 and molecular marker 2; the molecular marker 1 is located at the 2552nd base of the buffalo genus reference gene sequence NC_049568 or the 2540th base of the Bos genus reference gene sequence AB074968, and the polymorphism is T or C; the molecular marker 2 is located at the 2555th base of the buffalo genus reference gene sequence NC_049568 or the 2543rd base of the Bos genus reference gene sequence AB074968, and the polymorphism is G or A. The molecular marker combination of the present invention can be effectively used for detecting adulterated Bos genus milk in buffalo milk.

The invention discloses a noninvasive exosome mRNA BRAF V600E mutation Tag-PCR (Tag-Polymerase Chain Reaction) detection method, which comprises the following steps: separating an exosome sample from a body fluid; the method comprises the following steps: by taking mRNA as a template, adding an exogenous reference gene and a primer pair for detecting the exogenous reference gene, and meanwhile, carrying out a first round of PCR amplification reaction by using a Tag-ARMS primer pair for specifically enriching exosome mRNA BRAF gene V600E mutation so as to form a PCR product; a PCR product is used as a template, a system without UNG enzyme is used for amplification, a primer pair for detecting a reference gene is added, a Tag primer pair for specifically enriching and detecting V600E mutation of the human exosome mRNA BRAF gene and a probe sequence are used at the same time, and then a second round of PCR amplification reaction is carried out; and judging the mutation state of the V600E site of the BRAF gene of the mRNA of the exosome sample.

The invention discloses a brassica chinensis nitrate stress reference gene and primers and application thereof, and relates to the field of molecular biology, the reference gene is a Bc60S RPL4-1 gene, the nucleotide sequence of the Bc60S RPL4-1 gene is shown as SEQ ID NO: 2, and the nucleotide sequence of the Bc60S RPL4-1 gene is shown as SEQ ID NO: 3. A primer for amplifying the reference gene comprises a forward primer with a nucleotide sequence as shown in SEQ ID NO. 10 and a reverse primer with a nucleotide sequence as shown in SEQ ID NO. 11. The invention provides a reference gene Bc60S RPL4-1 gene which is highly stable in expression in brassica chinensis nitrate stress research, and the accuracy and reliability of quantitative analysis of brassica chinensis nitrate stress related gene expression are remarkably improved.

The application discloses a kit for detecting LINC01480 gene expression and a preparation method and application thereof, relates to the field of gene expression kits, and is used for detecting LINC01480 gene expression and comprises a specific primer pair for the LINC01480 gene, a specific primer pair for a reference gene GAPDH, a positive control lentivirus vector containing a LINC01480 gene segment, a negative control lentivirus vector and an RT-qPCR reaction system. The application non-invasively monitors MRD of MM patients after treatment by detecting LINC01480 gene expression, so as to improve the convenience and accuracy of index monitoring of MRD of the MM patients after treatment.

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 discloses a kit for detecting mRNA (messenger Ribonucleic Acid) in E6 / E7 regions of 14 high-risk human papilloma viruses, and relates to the technical field of pathogen detection, and the technical key points are as follows: the kit is used for detecting E6 / E7 mRNA of at least one high-risk HPV, the high-risk HPV comprises HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68 types, and the HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68 types can be used for detecting the E6 / E7 mRNA of at least one high-risk HPV. The kit comprises a primer probe combination for detecting the 14 high-risk HPVs and a primer probe combination for detecting the reference gene ACTB. By detecting HPV E6 / E7mRNA, the kit can provide higher specificity than traditional DNA detection and reduce false negative results, so that the accuracy of cervical cancer screening is improved, the kit can early recognize high-risk HPV infection, facilitates timely discovery of cervical cancer precancerous lesions and provides possibility for early intervention, and the kit can distinguish at least 14 high-risk HPVs and has good application prospects. Accurate virus genotype information is provided for clinic, and personalized treatment and prognosis evaluation are facilitated.

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.

The invention provides a primer probe group, a kit and a detection method for detecting diarrhea pathogens of children based on multiple PCR (Polymerase Chain Reaction) combined with time-of-flight mass spectrometry. The primer probe group comprises specific primers and probes aiming at 10 diarrhea pathogens (5 bacteria and 5 viruses) and a human-derived reference gene. Specific primers and probes are designed in a pathogen gene conserved region, targeted detection is achieved, and specificity is high; 10 diarrhea-causing pathogens and a human-derived reference gene in an excrement sample can be simultaneously detected in the same PCR reaction system and the single-base extension system, the sensitivity and the accuracy are high, and the detection flux is improved by combining a full-automatic chip spotting instrument; the cost is low, the detection period is short, and reliable technical support is provided for diagnosis and treatment, epidemic situation monitoring and infection control of children with infectious diarrhea.

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.