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. 2020 Sep 16;10:461. doi: 10.3389/fcimb.2020.00461

Figure 1.

Figure 1

CtpFCKD strain is attenuated for growth in macrophages. (A) Screening of CtpFCKD colonies. Quantitative RT-PCR showing downregulation of ctpF transcript with respect to levels in Mtb H37Rv, in putative knockdown colonies with sgRNA1, and sgRNA2. (B) In vitro growth curve of Mtb H37Rv (pink), dcas9-pRH2521 (orange), and ctpFCKD (olive green) in 7H9-ADC media. Graph depicts fold change in ctpF transcript levels in Mtb H37Rv in late log and stationary phase as compared to early log phase (C) and in different stresses as compared to mid-log phase (D). 16s rRNA was used as internal control in (A,C,D). (E) Growth profile of Mtb H37Rv, dcas9-pRH2521, and ctpFCKD in THP-1 macrophages. Six hundred ng/ml ATc was added after every 48 h to maintain conditional knockdown in (A,B). The values shown are mean of three independent experiments and error bars represent standard deviation. *, ** and NS indicate p < 0.05; p < 0.01 and p-value Non-significant, respectively, as analyzed by unpaired student's t-test.