## 3 Results
### Introduction of SNPs and Gene Deletions into M. luteus and Evaluation of comEA/EC Promoter Activity  
In order to confirm that individual point mutations found in the genomes of M- and R-mutants are responsible for increased or, respectively, decreased _comEA/EC_ promoter activity as indicated by the observed blue/white phenotypes, we introduced single conspicuous mutations into the genome of non-EMS-treated _M. luteus_ trpE16. The markerless introduction of point mutations into the genome was performed using a _codB/codA_-based counter-selection system [34]. Because of the lack of transformability of the reporter strain _M. luteus_ trpE16 \(\Delta\)_comEA/EC:lacZ-Kan_, which lacks functional _comEA_ and _comEC_ genes, the genome editing had to be conducted in the trpE16 parental strain. After confirmation of successful SNP re-insertion by sequencing, each single-SNP-containing strain was transformed with a DNA fragment carrying the \(\Delta\)_comEA/EC:lacZ-Kan_ construct, leading to strains with single point mutations, which otherwise were isogenic to the original reporter strain _M. luteus_ trpE16 _AcomEA/EC:lacZ-Kan_. This allowed the comparison of the LacZ reporter enzyme activity between isogenic strains, differing only by selected point mutations or gene deletions. In total, nine SNPs belonging to four different clusters were re-inserted \begin{table}
\begin{tabular}{c c c c c c} \hline \hline
**Cluster** & **SNP** & **Mutant/s** & **Mut\_Type** & **Locus** &
\begin{tabular}{c} **ORF Ori-** \\ **entation** \\ \end{tabular} & \multicolumn{1}{c}{**Annotated Product**} \\ \hline
1 & G103107A & M89, M106, MT & Transition & \
\begin{tabular}{c} Mult\_00960 \\ \end{tabular} & \(>\) &
\begin{tabular}{c} Cof subfamily of IIB subfamily of \\ haloacid dehalogenase superfamily \\ Protein of unknown function DUF88 \\ \end{tabular} \\
1 & G106197A & M33 & Transition &
\begin{tabular}{c} Mult\_00990 \\ \end{tabular} & \(<\) & Amino acid transporter \\
3 & G341241A & M101, M1080, & Transition &
\begin{tabular}{c} Mult\_03135 \\ \end{tabular} & \(<\) & N6-DNA Methylase \\  & & M42 & & & \\
3 & G341448A & M96 & Transition &
\begin{tabular}{c} Mult\_03135 \\ \end{tabular} & \(<\) & Hypothetical protein \\
4 & C575082T & M93 & Transition & - & - & Intragenic sequence \\
4 & G575822A & M28, M28B & Transition &
\begin{tabular}{c} Mult\_05280 \\ \end{tabular} & \(<\) & Hypothetical protein \\
7 & C1061676T & M89, M106, MT & Transition & \
\begin{tabular}{c} Mult\_09880 \\ \end{tabular} & \(>\) &
\begin{tabular}{c} ATP-dependent Clp protease \\ proteolytic subunit ClpP \\ ATP-dependent Clp protease \\ ATP-binding subunit ClpX \\ ATP-dependent Clp protease \\ ATP-binding subunit ClpX \\ ATP-dependent Clp protease \\ ATP-binding subunit ClpX \\ ATP-binding subunit ClpX \\ SAF domain-containing protein \\ Intergenic sequence (Mlut\_14660 \\ promoter) \\ Intergenic sequence (Mlut\_14660 \\ \end{tabular} \\
13 & G16077117A & M1090 & Transition & - & - &
\begin{tabular}{c} Intergenic sequence (Mlut\_14660 \\ promoter) \\ \end{tabular} \\
13 & C1607204T & M96 & Transition &
\begin{tabular}{c} Mult\_14660 \\ \end{tabular} & \(>\) & DNA-binding protein, excisionase \\  & & M9, M7, M43, & & & \\
13 & C1607261T & M47, M101, & Transition &
\begin{tabular}{c} Mult\_14660 \\ \end{tabular} & \(>\) & DNA-binding protein, excisionase \\  & & M44, M1080, & & & \\
13 & C1607314T & M93 & Transition & \
\begin{tabular}{c} Mult\_14660 \\ \end{tabular} & \(>\) &
\begin{tabular}{c} DNA-binding protein, excisionase \\ family \\ \end{tabular} \\
15 & C2060795T & M93 & Transition & \
\begin{tabular}{c} Mult\_19080 \\ \end{tabular} & \(<\) &
\begin{tabular}{c} Flavonodia \\ Thiol-disulfide isomerase such as \\ thioercoxin \\ Thiol-disulfide isomerase such as \\ thioercoxin \\ Thiol-disulfide isomerase such as \\ thioercoxin \\ DNA-binding protein, excisionase \\ family \\ RNA polymerase sigma factor, \\ sigma-70 family \\ \end{tabular} \\ \hline \hline \end{tabular}
\end{table}