Table 1 Conjugation results showing that the plasmid backbone has an effect on achieving successful editing
Construct | Total number of exconjugants | Percent of exconjugants edited and segregated on 1st patch |
|---|---|---|
pVZ321 | >250 | – |
pVZ321 + sSpcas9 | 0 | – |
pVZ321 + nblA sgRNA, ∆nblA editing template, sSpcas9 | 0 | N/A |
pVZ321 + nblA sgRNA, ∆nblA editing template, sSpcas9 (no promoter) | 0 | N/A |
pVZ321 + nblA sgRNA, ∆nblA editing template, sSpcas9 (no RBS) | 4 | 0/4 (0 %) |
pCRISPomyces-2 + nblA sgRNA, ∆nblA editing template, sSpcas9 | 21 | 16/16 (100 %) |
pCRISPomyces-2 + nblA sgRNA, ∆nblA repair template—sSpcas9 | 32 | 3/10 (30 %) |