Fig. 4

Phenotypic analysis of iRJ1 integrants harboring reporter plasmids. a Western immunoblot analysis to detect RNaseJ1-6xHis protein expressed in E. coli BL21(DE3) with integration of pBAD33-containing control DNA fragment (iBAD33, lane 1–3) and RNaseJ1-expressing gene cassette (iRJ1, lane 4–6) cultured in 0, 0.0012, and 0.0037% (w/v) arabinose as shown by the wedge above the lanes. Total protein-stained membrane is shown in the left panel and immunodetection of RNaseJ1-6xHis using Anti-6X His IgG, CF™680 (Sigma-Aldrich, Merck KGaA, Germany) is shown in the right panel. The expected size of RNaseJ1-6xHis is 63 kDa, as shown in the arrow. Lane M indicates KaledioscopeTM prestained protein ladder (Bio-Rad). b Growth analysis of iRJ1 integrants harboring reporter plasmids, pEGFP-hDHFR (pEGFP-hDHFR_iRJ1), pRzIEGFP-hDHFR (pRzI_iRJ1), and pRzIIEGFP-hDHFR (pRzII_iRJ1). OD₆₀₀ was measured every hour from 0 to 8 h cultivation time and data were plotted using the Growthcurve package in R software. Data are shown for each cell type grown in the presence or absence of 0.0012% (w/v) arabinose. Points represent the mean of 5 experiments and error bars represent 95% confidence intervals. P-values from two sample t-tests for pEGFP-hDHFR_iRJ1, pRzI_iRJ1, and pRzII_iRJ1 integrants comparing growth in the presence or absence of 0.0012% (w/v) arabinose are 0.00427, 0.00523, and 0.00373, respectively. c Fluorescence intensity of pEGFP-hDHFR_iRJ1, pRzI_iRJ1, and pRzII_iRJ1 integrants cultured in 0.0012% (w/v) arabinose relative to untreated control. Box plots show relative fluorescence data distribution. Dots indicate relative fluorescence data from individual experiments, and median values are indicated by the bold black line. P-values from single value two-tailed t-tests comparing group mean to 1 of pEGFP-hDHFR_iRJ1, pRzI_iRJ1, and pRzII_iRJ1 are 0.005381, 0.04022, and 0.03996, respectively