阅读说明：本技术 识别细极链格孢菌的单克隆抗体及其杂交瘤细胞株AtD2 (Monoclonal antibody for identifying alternaria tenuis and hybridoma cell strain AtD2 thereof ) 是由 赵伟春 徐云飞 李吉二 王海林 于 2019-12-03 设计创作，主要内容包括：本发明公开了一种识别细极链格孢菌Alternaria tenuissima的单克隆抗体及其杂交瘤细胞株,所述识别细极链格孢菌的单克隆抗体,由保藏编号为：CCTCC No：C2019228的杂交瘤细胞株分泌产生；所述杂交瘤细胞株命名为杂交瘤细胞株AtD2,于2019年10月17日保藏于中国典型培养物保藏中心(CCTCC),保藏编号为：CCTCC No：C2019228。该单克隆抗体被用于由细极链格孢菌侵染引发的动植物病害的鉴定、动态监测以及细极链格孢菌的生物学研究,通过BaL b/c小鼠腹腔注射该细胞株,即可获得大量的该单克隆抗体,与多克隆抗体相比,具有纯度高,专一性强,重复性好等优点。(The invention discloses a monoclonal antibody for identifying Alternaria tenuissima and a hybridoma cell strain thereof, wherein the monoclonal antibody for identifying the Alternaria tenuissima is prepared from the following components in parts by weight: CCTCC No: c2019228 hybridoma cell strain secretion; the hybridoma cell strain is named as a hybridoma cell strain AtD2, is preserved in China Center for Type Culture Collection (CCTCC) in 2019, 10 and 17 months, and has the preservation number as follows: CCTCC No: C2019228. the monoclonal antibody is used for identification and dynamic monitoring of animal and plant diseases caused by Alternaria tenuissima infection and biological research of Alternaria tenuissima, and a large amount of monoclonal antibody can be obtained by injecting the cell strain into the abdominal cavity of a Balb/c mouse.)

1. A monoclonal antibody recognizing Alternaria tenuis, which is represented by the deposit number: CCTCC No: c2019228.

2. A hybridoma cell strain is named as a hybridoma cell strain AtD2 and is preserved in China Center for Type Culture Collection (CCTCC) in 2019, 10 months and 17 days, wherein the preservation number is as follows: CCTCC No: C2019228.

Technical Field

The invention belongs to the field of animals, and particularly relates to a monoclonal antibody for identifying alternaria tenuis and a hybridoma cell strain AtD2 thereof.

Background

Alternaria tenuissima, a fungus of Alternaria of the subdivision Deuteromycotina, is one of the pathogenic fungi of fritillaria melasma. The pathogenic bacteria can be lost in soil with the hypha to live through the winter, and the fritillaria is infected again in the next year. Alternaria tenuissima is also a pathogenic bacterium of black spot of numerous crops, and causes serious loss to agricultural production every year. The alternaria fungus is of various types, and the colony, hypha and spore forms of related species are similar, so that the alternaria fungus is difficult to distinguish by means of the characteristics. The monoclonal antibody (monoclonal antibody) combined enzyme-linked immunosorbent assay (ELISA) for alternaria tenuis disclosed by the invention has the characteristics of high sensitivity and strong specificity, and is suitable for detecting a large amount of alternaria tenuis in fields, so that a foundation is laid for the identification and biological research of the alternaria tenuis by applying the antibody and the dynamic monitoring of diseases infected by the alternaria tenuis, such as thunberg fritillary black spot and the like.

Disclosure of Invention

The invention provides a monoclonal antibody for identifying alternaria tenuis, which is prepared from the following components in percentage by weight: CCTCC No: c2019228.

The invention also provides a hybridoma cell strain which is named as hybridoma cell strain AtD2 and is preserved in China Center for Type Culture Collection (CCTCC) in 2019, 10 and 17 months, wherein the preservation numbers of Wuhan university in Wuhan City are as follows: CCTCC No: C2019228.

the invention has the following beneficial effects:

(1) the monoclonal antibody has strong specificity: the monoclonal antibody has strong reaction with Alternaria tenuissima antigens, does not react with Alternaria tenuissima, Fusarium oxysporum, Fusarium solani, Fusarium equiseti, Fusarium semitectum, Botrytis cinerea, Phoma stem rot fungus and phomopsis stolonifera antigens, and can be used for detecting Alternaria tenuissima and thunberg fritillary black spot caused by the Alternaria tenuissima.

(2) The detection sensitivity of the monoclonal antibody is high: the indirect ELISA detection result shows that the detection sensitivity of the monoclonal antibody to the antigen prepared by the alternaria tenuis hyphae and the spores is 12.21ng/mL (namely 1.221ng per well).

(3) The monoclonal antibody is IgM.

(4) The monoclonal antibody-bound target protein is single: the monoclonal antibody only binds to an antigen protein of about 43kDa of Alternaria tenuissima.

(5) The detection sensitivity of the fritillaria thunbergii extracting solution on the monoclonal antibody is not influenced: the monoclonal antibody has no cross reaction to the thunberg fritillary bulb protein extracting solution; after the thunberg fritillary bulb protein extracting solution is added into the antigen prepared from the alternaria tenuissima hypha and the spores, the detection sensitivity of the monoclonal antibody is not influenced by the thunberg fritillary bulb extracting solution, the content of the thunberg fritillary bulb protein extracting solution is 12.21ng/mL (namely 1.221ng per hole), and the method has good development and application prospects.

Drawings

FIG. 1 Titer test of AtD 2;

FIG. 2 reaction of AtD2 with different fungal antigens;

FIG. 3 detection sensitivity assay of AtD 2;

FIG. 4 type and subclass identification of AtD 2;

FIG. 5 detection of AtD2 bound target protein;

FIG. 6 influence of the Bulbus Fritillariae Thunbergii extract on the detection sensitivity of AtD 2.

Detailed Description

The invention is further explained below with reference to the figures and the examples.