Table 1 Culex quinquefasciatus cutting and transgenesis efficiencies of different HDR templates.

HR plasmid	Injected G0			Efficiency (per G0 germline)^a		Overall efficiency (out of total G1s)^b
HR plasmid	Injected eggs	Adult survivors	Survival	Cutting: cd−/cd− (%)	Transgenesis DsRed+ (%)	Cutting: cd−/cd− (%)	Transgenesis DsRed+ (%)
“Original”	815	105	12.88%	1/105 (0.95%)	0/105 (0%)	6/5377 (0.11%)	0/5377 (0%)
“Loop”	1480	55	3.72%	3/55 (5.45%) (p value = 0.055) ns	1/55 (1.82%) (p value = 0.167) ns	25/3148 (0.79%) (p value < 0.0001)****	4/3148 (0.13%) (p value = 0.014)*
“Loop + Mutation”	1151	24	2.09%	1/24 (4.17%) (p value = 0.188) ns	1/24 (4.17% (p value = 0.025)*	22/1126 (1.95%) (p value <0.0001)****	4/1126 (0.36%) (p value = 0.0008)***

A one-tail randomization test for a difference in proportions was performed to determine whether the Loop or the Loop + mutation scaffold variants caused an increase in transgenesis when compared to the “Original” gRNA; the obtained p values are reported in the table in bold type for these comparisons for either the “Efficiency per G0 germline” or “Overall efficiency (G1)” along with the significance expressed in using the asterisk convention.
ns not significant.
^aG0 germline cutting and transgenesis efficiencies were calculated as numbers of independent pools that produce either cd−/cd− mutant (cutting) or DsRed+ (transgenesis) animals, divided by the total number of crossed G0s. While each pool contains several G0 individuals, our calculations assume only one editing event happened in each positive pool and may underestimate the cutting and transgenesis rates.
^bThe overall cutting and transgenesis efficiencies were calculated as the number of G1 individuals with either cd−/cd− (cutting) or DsRed+ (transgenesis) phenotypes divided by the total number of G1s.

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