32 results about "Base-Base Mismatch" patented technology

The invention relates to a method for detecting single base mutation in the field of gene mutation detection. The method comprises the following steps: S1, designing a fluorescent signal probe according to the target gene; the fluorescent signal probe is completely complementary to the mutant gene of the target gene, and forms a single base mismatch with the wild-type gene of the target gene; S2, Add the sample to be tested of the target gene and λ exonuclease to the solution containing the fluorescent signal probe to form a reaction system; S3, obtain the real-time fluorescence curve of the reaction system during the reaction, and judge whether the sample to be tested of the target gene is There is a single base mutation. The method of the invention can quickly and accurately detect the single-base mutation gene sequence, has high selectivity for the single-base mutation gene sequence, and can be used for low-abundance mutation detection. In addition, the detection cost of the method is low, easy to prepare, easy to operate, high in repeatability, and easy to be extended to non-professionals for operation.

Amino acid residue misincorporations are necessarily found in sequence variants at low concentrations in admixture with expressed polypeptides, resulting from one or more base mismatches within codons susceptible to amino acid residue misincorporation during transcription and / or translation. The invention provides a method of optimizing the coding sequences of a polynucleotide that encodes a polypeptide, wherein at least one codon is susceptible to amino acid residue misincorporation. The method of the invention can be used to reverse-engineer an unknown coding sequence, which encodes the same polypeptide, but differs in said at least one codon from the known coding sequence. The method can further be used to alter the immunogenic potential of an expressed polypeptide. Thus, the invention is useful in engineering optimized polynucleotides encoding polypeptides.

The invention belongs to the field of recognition of single base mismatches of nucleic acid molecules, and discloses a method for simultaneously distinguishing single base mismatches at the single molecule level. The method for simultaneously distinguishing single base mismatches at the single-molecule level includes: combining nanopore electrochemical technology to determine a non-labeled co-arm-type improved molecular beacon; Single-molecule strategies simultaneously discriminate between perfectly complementary and single-base mismatched sequences. The present invention combines nanopore electrochemical technology to design a non-marked co-arm type improved molecular beacon, which shows a higher signal-to-background ratio (~104), higher thermodynamic stability and faster hybridization rate. This label-free single-molecule strategy realizes the simultaneous distinction of fully complementary and single-base mismatched sequences at the single-molecule level, and the invention has potential application value in the fields of rapid detection, accurate identification, and early clinical diagnosis.

The invention discloses a detection method for the visual recognition of microRNA (Ribonucleic Acid) on the basis of G-quadruplex probe structure unwinding. A signal attenuation method constructed bycombining catalytic characteristic of a G-quadruplex combined with heme with the structure change of the G-quadruplex can distinguish miRNAs of single-base mismatch and multibase mismatch, and also can distinguish different miRNA family members. On the basis of signal attenuation, sensitivity still can be 4.5nmol / L. The probe used by the invention is simple in design, and does not relate to a complex reaction process. The detection method has an important meaning for the detection of miRNA.

The invention discloses a method and application for rapidly detecting nucleic acid based on electrochemical potential pretreatment technology. In the method, the PBS buffer is used as the electrolyte, the probe and the target nucleic acid fragment are added into the PBS buffer, and a nucleic acid hybridization solution is formed according to the principle of complementary base pairing. The sensing electrode is subjected to electrochemical pretreatment in a potential range greater than 1.8V, and then SWV is used to selectively detect target nucleic acids. The nucleic acid detection method of the electrochemical pretreatment technology is applied in the fields of disease characteristic miRNA-related electrochemical biosensing detection, medical diagnosis, and pathogen detection in food and environment. When this detection method is applied to nucleic acid, it has the following characteristics: high selectivity, which can detect single base mismatches for nucleic acid molecules; high sensitivity, with a detection sensitivity of 5×10 ‑18 M; the test results can be completely repeated; the operation process is economical and simple, there is no chain reaction, no amplification, no fluorescent indicator.

The invention relates to a single base mutation detection method combining nucleic acid temporary hybridizing and magnetic separating. The method comprises the following steps: A, designing a capturing probe and a signal probe according to a target gene; B, preparing a magnetic sphere solution from a magnetic sphere coupled with the capturing probe, adding the signal probe to conduct nucleic acid hybridization with a to-be-detected sample; C, conducting magnetic separation after finishing the nucleic acid hybridization, and then re-suspending the magnetic sphere in a low-metal ion solution; D, conducting magnetic separation again, detecting an obtained liquid supernatant, judging whether single base mutation exists in the to-be-detected sample, wherein the capturing probe is completely complementary with a non-mutation part of the target gene; the signal probe is completely complementary with a mutant type gene of the target gene, and forms a single base mismatch with a wild type gene of the target gene. The detection method is fast, and can be used in low-abundance mutation detection.

The disclosure relates to recombinant protein expression systems comprising genetically modified cells wherein the cells are transformed or transfected with tRNA genes to reduce base mismatch due to genetic degeneracy in the genetic code.

The invention provides a single-base mismatched target site DNA library specific to a single sgRNA molecule, and a carrier library generated by connecting the DNA library with a carrier. The invention also provides an application of the DNA library / carrier library in the study on the detection of sgRNA off-target effect.

The present invention proposes a DNA probe for enriching low-frequency DNA mutations. The DNA probe is complementary to the sequence containing the low-frequency DNA mutation and has LNA modification. The probe according to the embodiment of the present invention is complementary to a sequence containing low-frequency DNA mutations, and the probe has LNA modification. The inventors found that when the probe is complementary to a sequence containing low-frequency DNA mutations, if the low-frequency DNA mutations actually exist , which is completely complementary to the base of the DNA probe, its Tm value will be higher than the Tm value of the base mismatch between the wild-type DNA and the DNA probe, and it will be higher than that of the normal base without LNA modification. The Tm value increases more, and at an appropriate temperature, the DNA probe binds more to DNA containing low-frequency mutations and less to wild-type DNA, so that low-frequency DNA mutations can be specifically enriched.

The invention discloses a method used for detecting serum miRNA of patients with cancer based on short nucleotide chain connection. According to the method, specific recognition and cutting of unpaired single strand nucleic acids and double-stranded nucleic acids with mismatched bases are carried out, mismatching and partial complementation of hybridization of a nucleic acid probe with target miRNA are detected, fluorescence is generated via enzymatic reaction of horse radish peroxidase combined on the nucleic acid probe with a chemiluminescent substrate, rapid amplification of detection signals is realized, and the target miRNA content is determined based on fluorescence intensity. The detection results are accurate; operation is simple and convenient; cost is low; detection throughput is high; and nucleic acid extraction or amplification is not needed.