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. 2020 Jul 15;184(2):1004–1023. doi: 10.1104/pp.19.01420

Figure 7.

Figure 7.

A repaired T. levigata nfp2 pseudogene can replace PanNFP2 for nodule formation. A, Schematic representation of the NFP2 coding region with indicated replacements to avoid CRISPR targeting of inserted NFP2 genes of P. andersonii (PanNFP2cr) and a repaired T. levigata (TleNFP2cr). Blue arrows indicate guide RNA target sites, and red lines indicate T. levigata mutations. Region 1, Replacement of six codons at the sg1 site; region 2, replacement of five codons at the sg3 site plus repair of the T. levigata indel (red line); region 3, repair of the double stop codon in T. levigata (red line, black asterisks). The replacement of five codons at the sg2 site is not shown. B, PanNFP2cr and repaired TleNFP2cr can restore nodulation in the Pannfp2 mutant line C3 when driven by the PanNFP2 promoter, whereas PanNFP1 cannot. Nodulation was scored at 5 wpi with M. plurifarium BOR2. Error bars represent the sd. Asteriks indicate statistical significance by Mann-Whitney-Wilcoxon test (*P < 0.05 and ****P < 0.00001). C and D, Nodule and section of pNFP2:PanNFP2cr line 1. E and F, Nodule and section of pNFP2:TleNFP2cr line 1. Bars = 2 mm (C and E) and 100 μm (D and F).