8 results about "Lambda exonuclease" patented technology

The invention discloses a preparation method of light-regulated single-stranded DNA and application of the light-regulated single-stranded DNA in CRISPR nucleic acid detection. According to the invention, Lambda excision enzyme and a primer modified with PC-Linker are introduced into a detection system based on RPA-CRISPR / Cas12a, and a one-tube nucleic acid detection system without PAM limitation is constructed. By introducing Lambda excision enzyme, the strict dependence of the traditional CRISPR / Cas12a detection technology on PAM sites is broken through, and the application range of the CRISPR / Cas12a detection technology is remarkably widened. And the system is integrated into one-tube detection, so that the risk of aerosol pollution caused by opening a cover for multiple times and transferring a sample is avoided while the experimental operation process is simplified. According to the research, CRISPR / Cas12a and CRISPR / Cas13a are further integrated into a single-tube reaction system, so that the probability of false negative results is reduced in a double-gene detection manner, and the detection accuracy is improved.

A genome-wide R-loop detection method, in particular, a genome-wide R-loop detection method independent of affinity enrichment and a use. According to the genome-wide R-loop detection method, nucleases are used to perform enzyme digestion on genomic DNA fragments, and R-loop detection can be achieved by performing one-step enzyme digestion simply using a system of Nuclease P1, T5 exonuclease, and Lambda exonuclease. The whole R-loop detection process does not require affinity enrichment or labeling treatment. The method is not only simple and easy to operate, but also can avoid the loss of samples and fragments, thereby reducing the cost and risk of detection, and improving the applicability.

The application discloses a homogeneous fluorescence quantitative detection method for base deletion sites in genomic DNA sequences, and covalently labels known sequence oligonucleotide chains to the base deletion sites through a covalent connection reaction, and realizes quantitative detection of the base deletion sites in the genomic DNA sequences by using Lambda exonuclease and corresponding fluorescence probes. The application is simple in operation, high in sensitivity, small in required biological sample amount, and can conveniently, quickly, sensitively and accurately determine the content of the base deletion sites in biological samples, and has high application value in biomedical research, clinical diagnosis and drug development evaluation related to DNA damage and repair.

The application discloses a double-stranded nucleic acid detection composition based on lambda exonuclease and deoxyribozyme and application thereof, and belongs to the technical field of biological technology, and aims to efficiently and sensitively detect double-stranded nucleic acid. The double-stranded nucleic acid detection composition based on lambda exonuclease and deoxyribozyme comprises a double-stranded nucleic acid detection probe, lambda exonuclease and a reporter molecule, the double-stranded nucleic acid detection probe comprises one long single-stranded DNA named recognition chain R and n short single-stranded DNAs named Dz; the recognition chain R comprises a target binding domain and n Dz binding domains arranged in sequence from 5' to 3' in order; the 5' end of the recognition chain R is modified with a functional group; the Dz is a deoxyribozyme capable of cleaving the reporter molecule when the Dz is single-stranded, and the 5' end of the Dz is modified with sulfur. The application has high sensitivity and specificity, and the detection limit can be as low as a femtomole.

The invention discloses a method for replacing high-temperature denaturation to generate a single chain in multiple PCR (Polymerase Chain Reaction) detection reaction, a method for improving sensitivity and accuracy of liquid-phase hybridization detection of multiple PCR products and a matched kit. A phosphate group is designed and modified at the 5'end of a primer, so that a PCR product of which one end of one chain is phosphorylated is amplified; carrying out digestion treatment on a PCR amplification product by using Lambda exonuclease, and completely removing a DNA chain with phosphorylation at one end to obtain a single chain; the method disclosed by the invention replaces a common method for generating a single chain by high-temperature denaturation of a multiplex PCR detection reaction, eliminates the negative influence of DNA renaturation on hybridization of the probe and the target sequence, improves the hybridization efficiency, and improves the accuracy and sensitivity of a liquid chip technology platform in molecular detection application.

The present invention belongs to the field of biological analysis and relates to a gene detection kit for the acute leukemia gene Pax‑5a based on selective digestion of lambda exonuclease. When the target gene is present, the tail sequence of HP‑HCR forms a flat end with the Pax‑5a gene. After adding lambda exonuclease, the hairpin is gradually digested from the 5' end, and the Pax‑5a sequence is released from HP‑HCR and combined with the next hairpin probe to achieve the recycling of the gene and achieve the purpose of the first step of amplification. At the same time, HP‑HCR is cleaved into a single-stranded trigger chain, triggering a hybridization chain reaction. After the H1 chain is opened by the trigger chain, it hybridizes with the H2 chain sequence. The generated hybrid chain continues to bind to new H1 and H2 chains, and the fluorescent groups and quenching groups on H1 and H2 are away from each other, and the fluorescent signal is restored to achieve the second amplification of the Pax‑5a gene. Thereby achieving the detection of the acute leukemia Pax‑5a gene.

A method, which synthesizes closed circular single-stranded and double-stranded DNA molecules using in vitro enzymatic systems, is described. Circular single-stranded DNA molecules and double-stranded DNA molecules (e.g., relaxed, or supercoiled) with various sizes can be synthesized. Unwanted DNA molecules, e.g., unligated oligomers, can be removed by exonucleases, such as T5 exonuclease, T7 exonuclease, lambda exonuclease, E. coli exonuclease I and / or III. A method of converting the single-stranded circular DNA molecules into double-stranded circular DNA molecules is also described. The single-stranded and double-stranded circular DNA molecules can be used in a variety of applications.

A method, which synthesizes closed circular single-stranded and double-stranded DNA molecules using in vitro enzymatic systems, is described. Circular single-stranded DNA molecules and double-stranded DNA molecules (e.g., relaxed, or supercoiled) with various sizes can be synthesized. Unwanted DNA molecules, e.g., unligated oligomers, can be removed by exonucleases, such as T5 exonuclease, T7 exonuclease, lambda exonuclease, E. coli exonuclease I and / or III. A method of converting the single-stranded circular DNA molecules into double-stranded circular DNA molecules is also described. The single-stranded and double-stranded circular DNA molecules can be used in a variety of applications.