An identification method and application of two dna2 components of bbtv
An identification method and DNA2-R technology, applied in the field of identification of two DNA2 components of BBTV
Active Publication Date: 2019-08-23
INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI
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Although it has been reported at home and abroad that recombination can occur between DNA2 and DNA2, and between DNA2 and other components of BBTV, there is still no good identification in the prior art of whether these two DNA2 components recombine and the proportion of their offspring produced after recombination. how many ratios
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[0041] The embodiment of the present invention detects the two DNA2 components of the banana bunchy top virus in the infected BBTV banana leaf sample collected at random, and the specific detection steps include:
[0042] 1. Extract the total DNA in the banana leaf sample infected with BBTV, and use the CTAB method to extract, specifically:
[0043] (1) get 0.2g to infect BBTV banana leaf and grind into powder in liquid nitrogen;
[0044] (2) Add 1 mL of preheated 2×CTAB buffer, mix well, and incubate at 65°C for 10 min;
[0045] (3) Add 500uL chloroform / isoamyl alcohol (volume ratio, 24:1), mix well and centrifuge at 4°C for 10min;
[0046] (4) Take the supernatant, add an equal volume of chloroform / isoamyl alcohol (24:1), mix well and centrifuge at 4°C for 10 min;
[0047] (5) Take the supernatant, add pre-cooled 2 times the volume of absolute ethanol, and place it at -20°C for 30 minutes;
[0048] (6) Centrifuge at 4°C for 15 minutes, discard the supernatant, dissolve th...
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The invention provides an identification method of two DNA2 component of BBTV (banana bunchy top virus). The identification method comprises the following steps: 1) preparing a to-be-detected banana sample which is infected by BBTV, extracting total DNA and conducting PCR (polymerase chain reaction) amplification; 2) linking a PCR product to a plasmid vector, extracting plasmid and conducting NdeI enzyme digestion on the plasmid; and conducting electrophoresis detection to judge whether the sample contains two DNA2 components or not; and 3) conducting PCR amplification by taking the plasmid, which is cut into two strips in the step (2), as a template and conducting XhoII enzyme digestion; and conducting agarose gel electrophoresis detection so as to judge whether recombination occurs in the DNA2 or not and to judge a recombination rate in accordance with the enzyme digestion strips. According to the technical scheme provided by the invention, it can identify whether a banana bunchy top virus (BBTV) isolate contains the two DNA2 components or not, whether recombination occurs in the DNA2 components or not and can identify a method for calculating the recombination rate after the occurrence of the recombination; therefore, the invention lays a foundation for further deep research on the BBTV and for the prevention and control of the virus.
Description
technical field [0001] The invention belongs to the technical field of bioengineering, and in particular relates to an identification method and application of two DNA2 components of BBTV. Background technique [0002] According to the classification standard published by the International Committee on Taxonomy of Viruses in 2014, the Nanoviridae virus is one of the three DNA virus families that cause plant diseases, and the other two are Geminiviridae and Cauliflower Mosaic Virus. (Caulimoviridae). Banana bunchy top virus (BBTV) is one of the main members of the genus Babuvirus in the family Dwarfviridae. As a typical member of the genus, banana bunchy top virus can cause banana bunchy top disease (BBTD), which once brought huge economic losses to the banana industry in my country, and is also the main hazard to the development of the banana industry in the world. one. Electron microscope observation shows that the virus particles of banana bunchy top virus are equiaxed p...
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IPC IPC(8): C12Q1/70C12Q1/686
CPCC12Q1/686C12Q1/70C12Q2521/301
Inventor 余乃通刘志昕熊忠国周朋王健华张雨良张秀春
Owner INST OF TROPICAL BIOSCI & BIOTECH CHINESE ACADEMY OF TROPICAL AGRI SCI