15 results about "RpoB" patented technology

The invention belongs to the technical field of genetic engineering and enzyme engineering, and particularly relates to a genetically engineered bacterium for stably and efficiently producing L-isoleucine as well as a construction method and application of the genetically engineered bacterium. The genetically engineered bacterium takes E. coliTHRD as a host and contains an RNA (Ribonucleic Acid) polymerase beta-subunit mutant coding gene rpoBM; compared with a parent RNA polymerase beta-subunit, the RNA polymerase beta-subunit mutant has the advantage that the amino acid residue at the 618th site of the amino acid sequence is sequentially replaced with leucine from the N terminal to the C terminal. The genetically engineered bacterium disclosed by the invention can tolerate L-isoleucine and accumulate less alpha-ketobutyric acid, and has a very good industrial application prospect.

The invention discloses a chloroplast RNA edited gene OsPPR7 for temperature-sensitive regulation and control of rice and application of the gene OsPPR7, and belongs to the technical field of gene engineering. The CDS sequence of the OsPPR7 gene is as shown in SEQ ID NO. 1. The OsPPR7 is an RNA (Ribonucleic Acid) editing factor which has a bidirectional regulation and control function and is sensitive to temperature. Editing of multiple sites on the rpoB gene is negatively regulated and controlled, and meanwhile RNA editing of multiple genes such as ndhB and ndhD which are crucial to the chloroplast function is positively regulated and controlled under high-temperature stress. Under the stress of high temperature of 35 DEG C, the loss of functions can lead to obvious reduction of RNA editing efficiency of a plurality of ndh genes, and serious yellowing of seedlings is caused. The key function of the rice OsPPR7 gene in regulation and control of rice growth and development and high-temperature response is clarified for the first time, the importance of the OsPPR7 gene in maintenance of chloroplast functions under high-temperature stress is clarified, and a new gene resource and a technical path are provided for cultivation of new varieties of high-temperature-resistant rice.

The invention discloses a CRISPR-Cas13a (clustered regularly interspaced short palindromic repeats-CRISPR-Cas13a) double crRNA (ribonucleic acid) system which is established by using ERASE test paper and is used for detecting the rifampicin resistance of mycobacterium tuberculosis on site. The CRISPR-Cas13a double crRNA-system comprises a Cas13a protein and crRNA, or a complex formed by the Cas13a protein and the crRNA; the crRNA comprises an anchoring sequence and a guide sequence; the anchoring sequence is used for being combined with Cas13a protein; the guide sequence is used for targeting mutation target sequences of sites 531, 526, 516 and 513 in a mycobacterium tuberculosis drug-resistant gene rpoB gene; the target sequence of the mycobacterium tuberculosis is located at the 2317th site to the 2597th site of the rifampicin drug-resistant gene rpoB. On the basis of a CRSPR-Cas13a nucleic acid detection technology, result interpretation is performed through RAA amplification in combination with a line elimination method (ERASE) lateral flow chromatography test paper, and experiments prove that the kit can target high-frequency mutation sites of four rifampicin drug-resistant genotypes at the same time, has good detection sensitivity and specificity, and can be used for detecting the rifampicin drug-resistant genotypes of the rifampicin drug-resistant genotypes of the rifampicin drug-resistant genotypes of the rifampicin drug-resistant genotypes. The method can realize rapid, efficient, simple and convenient detection of mycobacterium tuberculosis rifampicin drug-resistant nucleic acid, has a sensitivity of 10 copies / [mu] L, and has an important application value.

The application provides a primer probe set and a kit for detecting a rifampicin-resistant gene of Mycobacterium tuberculosis, and belongs to the technical field of molecular biology detection.The primer probe set comprises RAA primers and qPCR primers targeting the rpoB gene, probe primers targeting the wild-type rpoB gene, and probe primers targeting the mutant rpoB gene.The application adopts the detection methods of RT-RAA and qPCR in sequence, and the above specific primer probe set can be combined to simultaneously and rapidly detect the 516, 526, 531 and 533 sites of the rpoB gene of Mycobacterium tuberculosis, has good specificity, and the detection sensitivity can reach 5 copies / ul, is 20 times higher than the sensitivity of qPCR (100 copies / ul), and has a 5% detection capability for heterogenic drug resistance mutation; the detection method can also greatly shorten the detection time.

The invention belongs to the technical field of microbiological detection, and particularly relates to a Klebsiella pneumoniae MLST typing primer group and method based on a hospital localized microbiological mNGS platform. The primer group comprises primer pairs for amplifying an rpoB gene, an infB gene, a pgi gene, a phoE gene, a gapA gene, an mdh gene and a tonB gene, and corresponding nucleotide sequences are as shown in SEQ ID NO: 1-14. According to the technical characteristics of short sequencing read length (50 bp) and single-ended sequencing of a next-generation sequencing platform in a hospital, the primer group for Klebsiella pneumoniae MLST housekeeping gene amplification is redesigned, and amplicons are adaptive to the sequencing working principle of the existing next-generation sequencing platform in the hospital by optimizing the length and position of the amplicons. By utilizing the primer group, the kit or the method, the genetic relationship between hospital infection pathogenic bacteria can be quickly and accurately judged (within 24 hours), the timeliness and scientificity of infection prevention and control are remarkably improved, and the primer group, the kit or the method have a wide clinical application prospect.

The invention relates to the technical field of bacterial detection, in particular to a primer group for simultaneously detecting sepsis-related pathogenic bacteria genes and drug-resistant genes and a matched method and device. The pathogenic bacteria genes are a Hafnia alvei rpoB pathogenic gene and an enterococcus faecium NusG pathogenic gene, and the drug-resistant genes are an oxazolidinone antibiotic drug-resistant gene optrA and a polymyxin antibiotic drug-resistant gene mcr-1. The primer composition comprises a nucleotide sequence as shown in SEQ ID No. 1 to SEQ ID No. 16. The primer composition provided by the invention can be used for simultaneously detecting two pathogenic bacteria pathogenic genes and two drug-resistant genes causing sepsis, and the primer group has high specificity and high sensitivity and can be used for accurately detecting the pathogenic genes in the same reaction system. The kit can realize simple, rapid and accurate detection, is suitable for on-site rapid detection, and can provide a new technical means for sepsis detection.

The invention discloses a crRNA combination for detecting mycobacterium tuberculosis and a drug-resistant mutation site thereof, a kit and application, and belongs to the technical field of molecular detection. The crRNA combination comprises a crRNA-MTB (Complementary Ribonucleic Acid-Methyl Terminals B) used for detecting the mycobacterium tuberculosis, a crRNA-MTB-rpoB used for detecting the rifampicin drug-resistant mutation site of the mycobacterium tuberculosis, and a crRNA-MTB-katG used for detecting the isoniazide drug-resistant mutation site of the mycobacterium tuberculosis; by designing the specific crRNA combination for detecting the mycobacterium tuberculosis and the drug-resistant mutation site thereof, the sensitivity of detecting the mycobacterium tuberculosis and the drug-resistant mutation site thereof can be remarkably improved, the detection time can be shortened, and synchronous detection of the mycobacterium tuberculosis and the drug-resistant mutation site thereof can be realized, so that the kit has the advantages that the detection sensitivity is greatly improved, and the detection time is shortened. The method has a good application prospect in detection of the mycobacterium tuberculosis and the drug-resistant mutation site of the mycobacterium tuberculosis.

The invention discloses a mycobacterium tuberculosis rpoB gene mutation detection method based on a CRISPR-Cas12a technology and a kit, and relates to the technical field of molecular biological detection. According to the present invention, through the design and the screening of the specific crRNA, the 531-crRNA4 with the nucleotide sequence represented by SEQ ID NO. 4 is successfully obtained; the sequence is combined with CRISPR-Cas12a and a PCR (Polymerase Chain Reaction) technology, can efficiently detect the rifampicin drug-resistant mutation site S531L, and has the remarkable advantages of high sensitivity, strong specificity, simplicity and convenience in operation and the like. The detection result of a clinical sample is highly consistent with the Sanger sequencing result, the detection time is shortened to be within 2 hours, and the method is suitable for rapid screening of clinical drug resistance. According to the technology, the time for doctors and patients to obtain detection reports is remarkably shortened, and the medical efficiency is effectively improved.

This invention belongs to the field of biodetection technology and discloses a method and application for analyzing antiviral and proviral bacteria based on metatranscriptome sequencing. Based on metatranscriptome sequencing, this invention utilizes different viral and bacterial marker genes to analyze and calculate the RPM values ​​of viral and bacterial species. Using the RPM values ​​of viruses and bacteria, Spearman correlation analysis is performed to obtain a correlation table showing the interactions between all bacteria and viruses. This invention overcomes the problem of incomplete bacterial genomes in metatranscriptome sequencing, enabling the detection of all viruses and bacteria in a single mosquito sample. The antiviral and proviral effects of bacteria are detected by the correlation between bacterial rpoB genes and viral RdRp load. Furthermore, the large mosquito sample size involved in this invention enhances the statistical reliability and ecological universality of the study, resulting in highly representative research results.

A strain of bacteria, HITLi7, based on the quantification of conserved gene abundance. T A rapid and specific detection method is involved, which uses the determination of the abundance of species-specific conserved genes to quickly detect HITLi7. T This invention addresses the challenges of complex microbial composition and the limited availability of HITLi7 strains in current bio-enhanced activated carbon processes. T The technical challenge of rapidly and specifically identifying and detecting abundance changes in indigenous bacteria presents a problem. This invention utilizes the double-positive reaction of the bispecific conserved genes gyrB and rpoB to further enhance the specificity of the detection results. The high sensitivity and accuracy based on qPCR technology further strengthen the reliability of the results. The detection method of this invention provides highly targeted results, avoiding the large amount of redundant data and complex processing procedures associated with high-throughput technologies. Compared to detection methods based on traditional bacterial morphology and physicochemical properties, it has wider applicability, higher sensitivity, and greater accuracy.

The application belongs to the technical field of genetic engineering and enzyme engineering, and particularly relates to a genetically engineered bacterium for stably and efficiently producing L-isoleucine as well as a construction method and application thereof. rpoB M The genetically engineered bacterium takes E. coli THRD as a host and contains a gene encoding an RNA polymerase beta-subunit mutant The amino acid sequence of the RNA polymerase beta-subunit mutant is sequentially replaced by leucine at the 618th amino acid residue from the N terminal to the C terminal direction compared with the parent RNA polymerase beta-subunit. The genetically engineered bacterium has good industrial application prospects and can tolerate L-isoleucine and accumulate less alpha-ketobutyric acid.

The present invention belongs to the field of genetic engineering technology, specifically relating to a mutant of the RNA polymerase β subunit and its application. The present invention obtains an RpoB mutant through gene editing technology. By changing the 560th amino acid of RpoB from proline to leucine, the strain significantly increases the number of viable cells during fermentation and improves its tolerance to the late-stage fermentation tank environment. This mutant is then applied to Escherichia coli producing human milk oligosaccharides, ultimately significantly increasing human milk oligosaccharide production.

The invention belongs to the technical field of immunological detection, and particularly relates to a primer probe composition for detecting drug resistance of mycobacterium tuberculosis complex rifampicin and isoniazide and application of the primer probe composition. A multicolor melting curve method is adopted, rifampicin drug-resistant gene mutation detection is carried out by detecting mutation of drug-resistant determining regions of 507-533 sites of a mycobacterium tuberculosis complex rpoB gene, and isoniazide drug-resistant gene mutation detection is carried out by detecting mutation of ahpC promoter regions, inhA promoter regions and katG315 codons. The drug resistance condition of the detected strain can be judged according to the Tm value of the melting curve. The result of the embodiment shows that the rifampicin drug resistance detection and the isoniazide drug resistance detection realize the detection of mutation of one drug in one tube without separating two tubes, the drug resistance condition of one drug can be independently detected or the drug resistance conditions of two drugs can be detected by the two tubes together according to different conditions during use, the application is flexible, and the cost is low. The application market is wider.

A combination of blockers for detecting drug resistance of Mycobacterium tuberculosis, including a first peptide nucleic acid (PNA) blocker, a second PNA blocker, a third PNA blocker, and a fourth PNA blocker. The first PNA blocker covers codons 511 and 513 of rpoB gene. The second PNA blocker covers codon 516 of rpoB gene. The third PNA blocker covers codon 526 of rpoB gene. The fourth PNA blocker covers codons 531 and 533 of rpoB gene. The sequences of the first, second, third, and fourth PNA blockers respectively match the wild-type sequences of the corresponding regions of rpoB gene. A kit and method for detecting drug resistance of Mycobacterium tuberculosis are also provided.

The invention relates to a primer group for detecting drug resistance of mycobacterium tuberculosis and rifampicin, a kit and application. The primer group disclosed by the invention comprises multiple PCR (Polymerase Chain Reaction) primers aiming at a mycobacterium tuberculosis IS6110 gene, an IS1081 gene and an rpoB gene, and the rpoB gene region comprises three amplification forward primer sequences corresponding to a 516 codon region, a 526 codon region and a 531 codon region, two locking sequences (LNA) corresponding to the amplification forward primer sequences and one amplification reverse primer sequence corresponding to the amplification forward primer sequences. The primer group can be used for quickly and accurately identifying the mycobacterium tuberculosis and identifying the rifampicin resistance of the mycobacterium tuberculosis; a result measured by the method is good in specificity and high in sensitivity, and can be directly read by naked eyes.