A genetically engineered single-chain antibody strain expressing resistance to citreogreen penicillin and its application
A yellow-green penicillin and single-chain antibody technology, applied in microorganism-based methods, anti-fungal/algal/lichen immunoglobulins, instruments, etc., can solve the problems of CIT toxins that have not been reported.
Active Publication Date: 2019-07-09
FUJIAN AGRI & FORESTRY UNIV
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Problems solved by technology
But so far, the research on the immunological detection method of CIT toxin based on genetically engineered antibody has not been reported.
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Embodiment 1
[0019] Example 1 Preparation and identification of the single-chain antibody of the present invention.
[0020] 1. Preparation of citreoviridin artificial complete antigen
[0021] In this experiment, citreoviridin (CIT) was cross-linked with carrier proteins BSA and KLH to form immunogenic artificial complete antigens CIT-BSA and CIT-KLH. The entire cross-linking process is carried out in two steps. In the first step, the hydroxyl group on the 3-position of the citreoviridin molecule is transformed into a carboxyl group by the succinic anhydride method; The amino groups are linked together through dehydration condensation. Specific steps are as follows:
[0022] (1) Weigh 60 mg of succinic anhydride powder and dissolve it in 500 μL of anhydrous pyridine, add an equal amount of 1 mol / L DMAP at the same time, preheat in a 65°C oven for 30-60 min, and activate succinic anhydride.
[0023] (2) Add the succinic anhydride solution directly to 1 mg of CIT solid, make it fully dis...
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Abstract
The invention belongs to the field of antibody engineering and in particular relates to a genetic engineering single-chain antibody strain for expressing anti-citreoviridin and application thereof. Man-made complete antigens CIT-BSA and CIT-KLH are synthesized through a succinic anhydride method and a carbodiimide two-step method; the genetic engineering single-chain antibody strain capable of stably secreting and expressing the anti-citreoviridin CIT is screened through utilizing a bacteriophage display technology; a single-chain antibody with functions is expressed and purified through constructing a prokaryotic expression vector and an ELISA (Enzyme-Linked Immuno Sorbent Assay) detection kit is assembled, so that qualitative and quantitative detection of residual CIT toxins in agricultural products is realized and a foundation is laid for commercially developing and producing high-quality enzyme-linked immuno sorbent assay detection kits and gold-labeled antibody detection cards.
Description
technical field [0001] The invention belongs to the field of antibody engineering, and specifically relates to a genetically engineered single-chain antibody strain expressing resistance to chrysogreen penicillin and its application. Background technique [0002] Citreoviridin (CIT) is a toxic small molecule mycotoxin, which is mainly secreted by Penicillum citreoviridin (PCV), widely exists in cereal crops and agricultural by-products, and mainly grows in low temperature and high humidity Harmful to human and animal health in such harsh environments. As a small molecular toxin, CIT has neurotoxicity, can inhibit the central nervous system, and can even cause death in severe cases; it also has cardiovascular toxicity, and many studies have shown that CIT is the main cause of cardiac beriberi; and Studies have reported that CIT is the suspected cause of Keshan disease; the toxin can inhibit the growth of Candida albicans and the activity of HIV-1 in vitro; many animal experi...
Claims
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IPC IPC(8): C12N1/21C07K16/14G01N33/53G01N33/52C12R1/21
CPCC07K16/14C07K2317/622G01N33/52G01N33/5308
Inventor 王荣智汪世华顾小松杨航钟燕芳凌素美
Owner FUJIAN AGRI & FORESTRY UNIV