35 results about "Methicillin resistance gene" patented technology

The invention discloses a method for eliminating mecA plasmids based on CRISPR/Cas9 technology. The method comprises: selecting an MRSA strain to perform PCR amplification to a DNA sequence of a C terminal of mecA gene coding transpeptidase; carrying out gel extraction for mecA genes; connecting the mecA genes with a T-pMD19 (simple) carrier and preparing DH5[alpha] competent cells; electro-transforming the DH5[alpha] competent cells through the obtained T-pMD19-mecA plasmids; extracting the T-pMD19-mecA plasmids and performing sequencing and verification; performing double digestion treatment for the T-pMD19-mecA plasmids and performing gel extraction to the mecA genes; connecting pET-21a (+) plasmids with the mecA genes; designing and synthesizing oligos; constructing pCas9 :: mecA plasmids; electro-transforming the obtained pET-21a (+)-mecA plasmids into an escherichia coli expression strain BL21 (D3); and electro-transforming the pCas9 :: mecA plasmids and the pCas9 plasmids into BL21 (D3)+pET-21a(+)-mecA competent bacteria and BL21 (D3)+pET-21a(+) competent bacteria. The method is simple to operate and good in specificity, and can effectively block spread of mecA to eliminating the mecA strain.

Disclosed are methods of identifying a methicillin-resistant Staphylococcus aureus (MRSA) or methicillin-sensitive Staphylococcus aureus (MSSA) in a sample. The present invention provides a diagnostic method comprising modification of sequences of S. aureus by converting non-methylated cytosine residues ultimately into thymidine residues in the target nucleic acid. The invention further provides for the detection of modified sequences derived from the spa gene, the mecA gene, and the integrated SCCmec cassette of S. aureus.

The invention relates to the field of microorganism detection, in particular to a staphylococcus aureus methicillin resistant strain PCR (Polymerase Chain Reaction) detection kit. The invention has the technical scheme of amplifying and detecting a highly conservative specific nucleic acid sequence nuc gene in the staphylococcus aureus gene by using the polymerase chain reaction and molecular beacon and fluorescent probe technologies and simultaneously amplifying and detecting the drug resistant nucleic acid sequence mecA gene to judge whether the strain is a methicillin resistant strain of staphylococcus aureus. The invention also provides PCR amplification primers of the nuc gene and the mecA gene and the sequence of the probe. The kit is applicable to the rapid detection of the staphylococcus aureus methicillin resistant strain in a clinical specimen of a suspect staphylococcus aureus infector and has the advantages of high sensitivity, high specificity, stability, timeliness, convenience for operation and the like.

The invention provides a primer and a probe for detecting drug resistance genes mecA in methicillin-resistant staphylococcus aureus (MRSA). The invention also provides a method and a detection kit for detecting drug resistance genes mecA in MRSA. According to the invention, the conserved sequence part of drug resistance genes mecA is directly detected without being affected by the mecA gene variation of strains, and a result is a gold standard of MRSA diagnosis. The specific primer and the probe provided by the invention are applicable to currently known strains containing sequence variation.

The invention relates to a double-chain small molecule interference nucleic acid for inhibiting and killing various drug tolerance bacteria using methicillin-resistant staphylococcus aurous as represents. The siNA of the invention is a double-chain molecule with 19 base pairs. A sense strand and a antisense strand respectively have two overhanging bases dT at 5' terminals, the GC content is 40-55; the aimed target sequence is selected from the genes relative with the vital movements of copy, transcription and translation in the staphylococcus aurous genome and an mecA gene correlative with the drug tolerance; said target sequence is preserved in 900f staphylococcus aurous; said target sequence is in a conservative sequence area in more than 900f staphylococcus aurous with distinct source with all the gene sequences in the human genome; the target sequence of the said siNA double-chain molecule is selected from SEQ ID NO. 1-325, the sense strand is a corresponding DNA or RNA sequence with the target sequence and the antisense strand is a corresponding RNA or DNA with the sense strand according to the complementary base law.

The invention discloses a reagent kit and method for quickly detecting staphylococcus MecA. The reagent kit comprises cell lysate, amplifying reaction liquid, colloidal gold detection liquid and an immunochromatography reagent strip. The reagent kit is characterized in that transcription products of MecA and 16S rRNA are obtained through NASBA amplification on a primer pair in the amplifying reaction liquid. The reagent kit for quickly detecting genes of the staphylococcus MecA is stable in detection result, high in accuracy rate, high in sensitivity, low in cost and short in time consumptionand suitable for clinical promotion. The detection method is suitable for clinically and quickly detecting methicillin resistant staphylococcus and screening staphylococcus at pharynx nasalis, is quick and accurate, is simple in equipment, is low in requirements for operators, and has potential in bedside detection and application in hospitals at different levels.

The present invention provides improved tests for the detection of methicillin-resistant Staphylococcus aureus bearing a variant mecA gene. The tests are particularly useful for eliminating certain false negative results due to the presence of this variant in MRSA in patient samples.

The invention provides a primer group and a method for detecting four kinds of bacteria by means of multiple polymerase chain reaction (PCR). The primer group comprises phoA gene primer pairs of escherichia coli, mdh gene primer pairs of klebsiella pneumoniae, femA gene primer pairs of staphylococcus aureus and mecA gene primer pairs of methicillin resistant staphylococcus aureus. A detection method comprises the steps of firstly, extracting a DNA target template; performing multiple PCR amplification on the extracted DNA target template by using the primer group; finally, detecting and analyzing the obtained multiple PCR amplification product by means of gel electrophoresis. According to the detection method, the four kinds of bacteria can be rapidly detected in the same reaction system; compared with the traditional unique PCR or blood culture detection method, the detection method provided by the invention has the advantages of time saving, simpleness and convenience.

The invention discloses a label-free fluorescence detection method of Staphylococcus aureus mecA gene and a detection kit, belonging to the analysis and detection field. A strategy in which clip probeHP is combined with exonuclease Exo III assisted target signal cascade recycle amplification is adopted. In G-quadruplex is adopted as a signal reporter molecule to detect mecA gene of Staphylococcusaureus without label. The method and the kit simplify the operation and reduces the cost. The whole detection process is characterized by rapid response, simple, free of mark and high sensitivity, and can be easily popularized without professional training. The detection method and the detection kit of the invention have important significance for the rapid detection of Staphylococcus aureus in the environment or food.

The invention discloses a primer and a probe composition and a kit. The primer and probe composition are used for detecting coagulase negative staphylococcus, Staphylococcus aureus and methicillin resistance genes; a coagulase negative staphylococcus detection target is a sodA gene, the staphylococcus aureus detection target is a femA gene, and the methicillin resistance gene is a mecA gene; The primer and probe composition include a forward primer, a reverse primer, and a probe for respectively detecting the sodA gene, the femA gene, and the mecA gene, which are shown as SEQ ID NO. 1 to SEQ ID NO. 9. The primer and a probe composition use real-time fluorescent quantitative PCR to detect the coagulase negative staphylococcus, staphylococcus aureus and methicillin resistance genes. The detection cycle is greatly shortened, the efficiency and the detection flux are improved, the detection requirement is catered for, and the specificity, sensitivity and accuracy are high, at the same time, no detection process is generated after amplification in the experiment, and the pollution problem in the experiment can be effectively solved.

The invention belongs to the field of clinical examination and diagnosis, and relates to a method and kit for rapidly detecting the drug resistance and virulence of staphylococcus aureus. The kit comprises three pairs of multiple PCR amplification primers, 2*PCR premix system, and sterilized double distilled water, wherein the multiple PCR amplification primers are spa gene upstream/downstream primers, mecA gene amplification upstream/downstream primers, and pvl gene amplification upstream/downstream primers. The kit comprises three pairs of PCR primers and thus can simultaneously amplify the spa, mecA, and pvl genes of staphylococcus aureus. The kit can be used to detect staphylococcus aureus, which is resistant to methicillin, in a clinical sample, can detect staphylococcus aureus with high virulence at the same time, and provides references for the clinical treatment on staphylococcus aureus infection. The invention establishes a method for rapidly detecting drug resistance and virulence of staphylococcus aureus, and the method is simple, rapid, and cheap and has an important meaning for the clinical treatment on staphylococcus aureus infection.

The invention belongs to the field of pharmacy, and relates to a benzene polyene sesquiterpene derivative and an application thereof in preparing an anti-resistant-bacteria medicine and preparation, and especially in preparing medicines and preparations used for inhibiting drug-resistant staphylococcus aureus. The benzene polyene sesquiterpene derivative provided by the invention comprises compounds 1 and 2, and is obtained by separation from a traditional Chinese medicine material asafoetida. As a result of test, the derivative has pharmacological activity against drug-resistant staphylococcus aureus. The compound 2 has a minimal inhibitory concentration (MIC) of 2mug/L against tetracycline-resistant staphylococcus aureus strain XU212 and methicillin-resistant staphylococcus aureus strain EMRSA-15, such that the activity is substantially better than control drugs tetracycline and oxacillin. For bacterium drug resistance, the compounds 1 and 2 show significant external pump inhibition effect against resistant staphylococcus aureus RN4220 with MsrA macrolide external pump protein and methicillin-resistant staphylococcus aureus EMRSA-16 with mecA gene. The derivative can be further used for preparing medicine preparations used for resisting external-pump drug-resistant staphylococcus aureus.

The invention relates to the technical field of biomedicine, and particularly relates to a system for simultaneously detecting methicillin-susceptible and methicillin-resistant staphylococcus aureus and a using method thereof. The system comprises an m-LAMP unit and a detection unit; the m-LAMP unit is used for performing loop-mediated isothermal amplification on femA gene and mecA gene simultaneously in the same reaction system; the m-LAMP unit comprises a first primer combination for performing loop-mediated isothermal amplification on the femA gene and a second primer combination for performing loop-mediated isothermal amplification on the mecA gene; and the detection unit is used for detecting femA gene and mecA gene products after loop-mediated isothermal amplification. The system canrealize simultaneous detection of MSSA and MRSA, and can be applied to practice operations of mechanisms such as hospitals and the like for detection of MSSA and MRSA, and effective reference is provided for formulating a timely and accurate treatment scheme.

The invention discloses primer sets for detecting an mecA gene of methicillin-resistant staphylococcus aureus on a severely-infected patient, and further relates to a method for detecting the methicillin-resistant staphylococcus aureus through the primer sets and a quick quantitative detection kit for detecting the methicillin-resistant staphylococcus aureus through the primer sets. According to the primer sets, the method and the kit, the best primer sequence and the best detection method for measuring the methicillin-resistant staphylococcus aureus with the mecA gene are determined, and a quick and convenient LAMP high-throughput detection method for laboratories is built.

The invention discloses an ultra-sensitive electrochemical LCR sensor for detecting methicillin-resistant staphylococcus aureus in synovial fluid. The ultra-sensitive electrochemical LCR sensor comprises the following steps of (1) designing two specific double-stranded short probes (S-dsDNA) according to conservative series of a MecA gene; (2) in an LCR reaction system, taking the MecA gene as a template, connecting two short series of probes through DNA ligase to form long double-stranded DNA (L-dsDNA), and then carrying out cyclic amplification by using an L-dsDNA template to form a large amount of L-dsDNA, and (3) fixing the formed L-dsDNA with sulfydryl and biotin modification on a BSA modified gold electrode through a gold-sulfur bond, dropwise adding avidin-PolyHRP to be combined with biotin on the L-dsDNA, finally putting the electrode into a base solution containing TMB and H2O2, and enabling H2O2 to oxidize TMB to generate an electrochemical signal under the catalysis of HRP. The sensor has the advantages of economy, quickness, high sensitivity, specificity and the like, can be used for detecting a single base mutation series, and realizes the detection of the MRSA in the synovial fluid of a patient infected around the joint prosthesis.

The invention discloses a pyrophosphoric acid detection kit for common pathogenic bacteria and drug-resistant genomes as well as a detection method and application of the pyrophosphoric acid detection kit. The kit is used for detecting 12 drug-resistant bacteria strains and drug-resistant genes in a sample to be detected, comprising genotypes of pseudomonas aeruginosa, acinetobacter baumannii, staphylococcus aureus, klebsiella pneumoniae, stenotrophomonas maltophilia, escherichia coli, enterococcus faecalis, enterococcus faecium, KPC gene, OXA-23 gene, SHV gene and MecA gene, and the kit comprises an amplification primer group, a sequencing primer group, an amplification solution, an amplification enzyme and a sequencing substrate. By adopting the pyrosequencing method, 12 pathogenic bacteria and drug-resistant genes with high pathogenicity and high detection rate can be detected at the same time, the pathogenic bacteria and the drug-resistant genes to be detected can be combined at will, detection of different frequencies and combinations can be carried out aiming at different environments, the detection process is flexible, simple and convenient, and the pyrosequencing result is rapid and readable.

A composition for diagnosing sepsis, and a method and a kit therefor. The kit includes a primer composition containing specific individual primers for amplifying: a segment of a 16s rRNA gene; a segment of an ITS gene; a segment of a Nuc gene; a segment of a MecA gene; segments of a VanA and a VanB; a segment of an invA gene; a segment of an ipaH gene; and a segment of a Cps gene. The kit also includes a probe composition containing specific individual probes for detecting: gram-negative bacteria; gram-positive bacteria; a Nuc gene; a MecA gene for checking whether or not MRSA has antibiotic resistance; a VanA and a VanB, for checking whether or not VRE have antibiotic resistance; and a gene for identifying a fungus.

The invention relates to the technical field of biological detection, in particular to primers, probes and a kit for multiplex PCR detection of seven drug-resistant genes. The kit disclosed by the invention can be used for carrying out joint detection on carbapenem enzyme genes KPC, NDM, IMP, VIM and OXA48 genes in carbapenem drug-resistant enterobacteriaceae bacteria, methicillin-resistant staphylococcus mecA genes and vanA genes in vancomycin-resistant multi-drug-resistant enterococcus in a rapid, accurate and ultrahigh-sensitivity manner; the change of the specific drug-resistant gene in the body of a blood flow infection patient can be rapidly monitored in real time, the condition of the patient is evaluated in time, and auxiliary treatment suggestions are provided for clinicians. The kit disclosed by the invention can also be used for active screening of drug-resistant genes of seven bacteria in CRE, MRSA and VRE of critically ill patients and immunosuppressive people without symptom colonization, and has extremely high application value in the field of scientific research and clinical application.

The invention discloses an application of 24-membered macrocyclic Schiff base in preparation of a drug for treating methicillin-resistant staphylococcus aureus infection. The 24-membered macrocyclic Schiff base is found to form a G-quadruplex structure with the core sequence of the MecA gene in the methicillin-resistant staphylococcus aureus through methods such as circular dichroism spectrum andWestern-Blot, so that the expression of the corresponding penicillin-binding protein PBP2a is inhibited. Pharmacological sensitivity experiments prove that the 24-membered macrocyclic Schiff base canreduce the minimum inhibitory concentration of methicillin-resistant staphylococcus aureus on oxacillin. When the use amount of the 24-membered macrocyclic Schiff base is 5 [mu] M, the minimum inhibitory concentration of methicillin-resistant staphylococcus aureus can be reduced to 0.5 [mu] g/mL to the minimum, and the effect of treating methicillin-resistant staphylococcus aureus infection by using beta-lactam drugs is achieved.