The invention discloses preparation of a chicken
eimeria tenella strain with EYFP knocked into an Et.
Actin gene. According to the invention, an FnCas12a
protein, crRNA synthesized
in vitro and a
homologous recombination fragment (an Et.
Actin gene containing a P2A-DHFR-EYFP
expression cassette) are introduced into chicken
eimeria tenella sporozoite by utilizing a Lonza 4D-Nucleofector nuclear transformation instrument, so that the editing of the chicken
eimeria tenella is realized. The edited
insect strain is screened through
pyrimethamine in a chicken body to obtain a high-purity
gene edited
insect strain. Then, the gene edited
insect strain is verified by utilizing PCR and
Sanger sequencing technologies, and it is proved that the EYFP successfully marks the Et.
Actin gene. After sporozoites of the strain are subjected to
cell culture and a chicken is infected,
cecum mucosa is taken and smeared to show that the EYFP is expressed in the whole
life history of the gene edited strain. The gene editing method can be used for performing fixed-point marking on a chicken
coccidiosis gene, and is beneficial to analyzing the function of the
coccidiosis gene. The invention also has important significance for research and development of novel anticoccidial drugs and vaccines.