96 Wells Fluorescence Reading And R Code Statistic For Analysis
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<strong>Overview</strong> Data points present in this dataset were obtained following the subsequent steps: To assess the secretion efficiency of the constructs, 96 colonies from the selection plates were evaluated using the workflow presented in Figure Workflow. We picked transformed colonies and cultured in 400 μL TAP medium for 7 days in Deep-well plates (Corning Axygen®, No.: PDW500CS, Thermo Fisher Scientific Inc., Waltham, MA), covered with Breathe-Easy® (Sigma-Aldrich®). Cultivation was performed on a rotary shaker, set to 150 rpm, under constant illumination (50 μmol photons/m<sup>2</sup>s). Then 100 μL sample were transferred clear bottom 96-well plate (Corning Costar, Tewksbury, MA, USA) and fluorescence was measured using an Infinite® M200 PRO plate reader (Tecan, Männedorf, Switzerland). Fluorescence was measured at excitation 575/9 nm and emission 608/20 nm. Supernatant samples were obtained by spinning Deep-well plates at 3000 × <em>g</em> for 10 min and transferring 100 μL from each well to the clear bottom 96-well plate (Corning Costar, Tewksbury, MA, USA), followed by fluorescence measurement. To compare the constructs, R Statistic version 3.3.3 was used to perform one-way ANOVA (with Tukey's test), and to test statistical hypotheses, the significance level was set at 0.05. Graphs were generated in RStudio v1.0.136. The codes are deposit herein. <strong>Info</strong> ANOVA_Turkey_Sub.R -> code for ANOVA analysis in R statistic 3.3.3 barplot_R.R -> code to generate bar plot in R statistic 3.3.3 boxplotv2.R -> code to generate boxplot in R statistic 3.3.3 pRFU_+_bk.csv -> relative supernatant mCherry fluorescence dataset of positive colonies, blanked with parental wild-type cc1690 cell of <em>Chlamydomonas reinhardtii</em> sup_+_bl.csv -> supernatant mCherry fluorescence dataset of positive colonies, blanked with parental wild-type cc1690 cell of <em>Chlamydomonas reinhardtii</em> sup_raw.csv -> supernatant mCherry fluorescence dataset of 96 colonies for each construct. who_+_bl2.csv -> whole culture mCherry fluorescence dataset of positive colonies, blanked with parental wild-type cc1690 cell of <em>Chlamydomonas reinhardtii</em> who_raw.csv -> whole culture mCherry fluorescence dataset of 96 colonies for each construct. who_+_Chlo.csv -> whole culture chlorophyll fluorescence dataset of 96 colonies for each construct. Anova_Output_Summary_Guide.pdf -> Explain the ANOVA files content ANOVA_pRFU_+_bk.doc -> ANOVA of relative supernatant mCherry fluorescence dataset of positive colonies, blanked with parental wild-type cc1690 cell of <em>Chlamydomonas reinhardtii</em> ANOVA_sup_+_bk.doc -> ANOVA of supernatant mCherry fluorescence dataset of positive colonies, blanked with parental wild-type cc1690 cell of <em>Chlamydomonas reinhardtii</em> ANOVA_who_+_bk.doc -> ANOVA of whole culture mCherry fluorescence dataset of positive colonies, blanked with parental wild-type cc1690 cell of <em>Chlamydomonas reinhardtii</em> ANOVA_Chlo.doc -> ANOVA of whole culture chlorophyll fluorescence of all constructs, plus average and standard deviation values. <strong>Consider citing our work. </strong> Molino JVD, de Carvalho JCM, Mayfield SP (2018) Comparison of secretory signal peptides for heterologous protein expression in microalgae: Expanding the secretion portfolio for Chlamydomonas reinhardtii. PLoS ONE 13(2): e0192433. https://doi.org/10.1371/journal. pone.0192433
