| Detecting DNA of causative agent of bovine moraxellosis in biological material of animals and feed, by PCR in real time having e.g. isolating DNA of agent of infectious disease from biological material of infected animal by sorption method | |
| 2023-06-13 | |
| 专利权人 | UNIV KUBAN STATE AGRARIAN (UYKU-Non-standard) |
| 申请日期 | 2023-06-13 |
| 专利号 | RU2023115553-A |
| 成果简介 | NOVELTY - Detecting DNA of the causative agent of bovine moraxellosis (Moraxella bovis) in biological material of animals and feed comprises PCR in real time including isolating DNA of the causative agent of infectious disease from biological material of infected animal using sorption method, and setting up one-step PCR with fluorescent detection with 40 cycles of real-time amplification using oligonucleotide primers, probes, fluorescent dyes, and internal control sample in the form of suspension of bacteriophage T4 with concentration of 5x 103specific to the region of the animal infectious disease pathogen's DNA genome phage particles per 1 μ l and positive control sample in the form of mixture containing in the same volume ratio fragments of the genomes of the pathogen of animal infectious disease and bacteriophage T4 with the nucleotide sequence of type 4 filaments at 5'-tacatataaatcacgcaaagc-3' as forward primer, T4R at 5'-tagtatggctaatcttattgg-3' as reverse primer. USE - The method is useful for detecting DNA of the causative agent of bovine moraxellosis (Moraxella bovis) in biological material of animals and feed. ADVANTAGE - None given. DETAILED DESCRIPTION - Detecting DNA of the causative agent of bovine moraxellosis (Moraxella bovis) in biological material of animals and feed comprises PCR in real time including isolating DNA of the causative agent of infectious disease from biological material of infected animal using sorption method, setting up one-step PCR with fluorescent detection with 40 cycles of real-time amplification using oligonucleotide primers, probes, fluorescent dyes, and internal control sample in the form of suspension of bacteriophage T4 with concentration of 5x 103specific to the region of the animal infectious disease pathogen's DNA genome phage particles per 1 μ l and positive control sample in the form of mixture containing in the same volume ratio fragments of the genomes of the pathogen of animal infectious disease and bacteriophage T4 with the nucleotide sequence of type 4 filaments at 5'-tacatataaatcacgcaaagc-3' as forward primer, T4R at 5'-tagtatggctaatcttattgg-3' as reverse primer, T4R at SU5-5'-acattggcactgaccgagttc-3'-black hole quencher 1 (BHQ1) as probe, measuring the accumulation of fluorescent signals in the channels of the corresponding fluorescent dyes for specific signal for testing the presence of infectious disease agent in animal and in the 6-carboxyfluorescein/green channel for internal control sample, and interpreting the results based on the presence or absence of intersection of the fluorescence curve with the threshold line, if the fluorescent signal accumulation curves extend up to the 35th cycle, then the reaction result is considered positive, and if the curves do not cross the threshold line or cross it after the 35-th cycle, then the reaction result is negative, where the DNA of the causative agent of bovine moraxellosis (Moraxella bovis) is isolated from the material of choice of smears from the mucous membranes of the conjunctiva and oropharynx, fragments of affected skin, organs, whole blood, milk, and sperm for the positive control sample, fragment of the genome of the causative agent of bovine moraxellosis (Moraxella bovis) with the nucleotide sequence (5'-3') is used as morab-F-gatttactcgatggtggtgc, morab-R-ccatcgagtgggtcatcgc, morab-PR6G-accgcttgtttggtggtaaaggcaac-BHQ1, and the fluorescent dye, hexachlorofluorescein/yellow is used for the DNA of the causative agent of Moraxella bovis. |
| IPC 分类号 | C12Q-001/68 |
| 国家 | 俄罗斯 |
| 专业领域 | 生物科学 |
| 语种 | 英语 |
| 成果类型 | 专利 |
| 文献类型 | 科技成果 |
| 条目标识符 | http://119.78.100.226:8889/handle/3KE4DYBR/21639 |
| 专题 | 中国科学院新疆生态与地理研究所 |
| 作者单位 | UNIV KUBAN STATE AGRARIAN (UYKU-Non-standard) |
| 推荐引用方式 GB/T 7714 | CHERNYKH O YU,BANNOV V A,DONNIK I M,et al. Detecting DNA of causative agent of bovine moraxellosis in biological material of animals and feed, by PCR in real time having e.g. isolating DNA of agent of infectious disease from biological material of infected animal by sorption method. RU2023115553-A[P]. 2023. |
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