Fig. 1

Construction of TgeIF-5A knockout mutant using CRISPR/Cas9. a Identification of the construction of pCRISPR-eIF-5A. (Lane M) DNA marker DL10000; (Lane 1) pSAG1-Cas9-U6-sg-eIF-5A digested by SalI enzymes; (Lane 2) pSAG1-Cas9-U6-sg-CDPK3 digested by Sal I enzymes. b Amplification of the 5′ and 3′ regions of eIF-5A and DHFR* cassette. (Lane M) DNA marker DL5000; (Lane 1) The amplification products of eIF-5A and CDPK3 5′ regions; (Lane 2) The amplification products of eIF-5A and CDPK3 3′ regions; 3: The amplification products of DHFR. c Identification of the homology template plasmid. (Lane M) DNA marker DL5000; (Lane 1) The amplification products of the 5′ region and DHFR fragment in each gene homologous template plasmid; (Lane 2) The amplification products of DHFR fragment and 3′ region in each gene homologous template plasmid; (Lane 3) The amplification products of the 5′ region, DHFR fragment and 3′ region in each gene homologous template plasmid. d Identification of the knockout mutant by PCR. (Lane M) DNA marker DL2000; (Lane 1) The amplification products of the 5′ region of each gene and part of the 5′ terminal of the DHFR fragment from gene editing or RH tachyzoites; (Lane 2) The amplification products of part of the 3′ terminal of the DHFR fragment and 3′ region of each gene from gene editing or RH tachyzoites; (Lane 3) The amplification products of part of the deleted genes containing the sgRNAs from gene editing or RH tachyzoites