Virus carrier particles and construction method and application thereof
A technology of virus vector and construction method, applied in the field of biomedicine, can solve the problems of no toxin effect, inability to inhibit stimulation well, and achieve the effect of reducing secretion
Active Publication Date: 2016-05-11
郑州可尔利尔生物科技有限公司
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Problems solved by technology
Rheumatologists have found Botox not only for tension headaches / migraines but injecting the toxin into the joints is also effective in RA (rheumatoid arthritis) and OA (osteoarthritis) patients who do not respond well to other medications However, because the endothelial cells and macrophages are not sensitive to toxins and do not have cell receptors for toxins, they cannot well inhibit the stimulation of inflammatory factors and inflammatory factors on sensory nerve cells, so Sensory neurons still secrete nociceptive transmitters
Method used
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Embodiment 1
[0047] A lentiviral vector particle includes pspax2, pmd2.G and pwpi plasmids. The pwpi plasmids include a promoter, a botulinum toxin type D enzyme active region and an internal ribosome entry site (IRES) sequence.
[0048] The synthetic botulinum toxin D-type enzyme active region gene is designed with a region for enhanced expression (ACCATGGGC), which is located before its initial coding, and contains PmeI sites for cloning, at the 5'and 3'ends, see the sequence table In the sequence 1. After botulinum toxin type D enzyme active region gene was digested and recovered by PmeI, it was cloned into pwpi plasmid after PmeI digestion and alkaline phosphatase treatment. The correct cloning direction was confirmed by PstI digestion and gene sequence determination.
[0049] The construction method of the above pwpi plasmid includes the following steps:
[0050] (1) First, 1μg of pwpi vector containing the marker gene (GFP or RFP) was digested with 1μLPME1, and digested in a 50μLeppendorf...
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The invention belongs to the technical field of biomedicine and particularly relates to virus carrier particles, a construction method and application thereof to preparation of medicine for treating human chronic pain. The virus carrier particles comprise pspax2 particles, pmd2.G particles and pwpi particles, and the pwpi particles comprise promoters, creotoxin enzyme activity areas and internal ribosome entry site sequences. The construction method of the virus carrier particles includes the steps that 1, a cell line lenti-x293T is cultured; 2, a DMEM culture medium is adopted; 3, the pspax2 particles, the pmd2.G particles and the pwpi particles are subjected to transfection into lenti-x293T cells; 4, transfection liquid is washed off, and culture liquid containing 10wt% calf serum is added for culturing; 5, the culture liquid is recycled, filtered and centrifuged, and the virus carrier particles high in drop degree can be obtained. The virus carrier particles are more targeted to treatment of chronic pain and can prevent vicious circle between a nervous system and peripheral cells.
Description
Technical field [0001] The invention belongs to the field of biomedicine technology, and specifically relates to a viral vector particle, a construction method thereof, and an application in a medicine for treating human chronic pain. Background technique [0002] Pain is an unpleasant feeling and emotional experience for actual or potential damage. It is a complex perceptual experience, including the response of nociceptors (pain receptors) that convey sensory information to stimuli such as location, type, and intensity. Pain and its treatment have always been regarded as an important and interesting field of medicine. [0003] Chronic pain is damage caused by tissues, often accompanied by long-term nerve damage. Chronic pain is caused by activation of nociceptors (pain receptors) leading to nociception (nerve processing harmful stimuli), and the sensation of pain begins with nerve afferents. The central system also plays a role, thereby maintaining the persistence of pain. [00...
Claims
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Patent Timeline
Login to View More IPC IPC(8): C12N15/867C12N15/861C12N15/864C12N15/869A61K45/06A61P29/00A61P19/02
CPCA61K45/06C12N15/86C12N2710/10043C12N2710/16043C12N2740/15043C12N2750/14143
Inventor 孟疆辉
Owner 郑州可尔利尔生物科技有限公司