New mutants of the thermotolerant strain Bacillus subtlis MSP4 were obtained by chemical mutagenesis with the use of ethidium bromide. The performed mutagenesis increased the maximum growth temperature relative to the parent strain from 50 to 60 °C. The next step will be to sequence the genomic DNA of the obtained mutants, and then carrying out a bioinformatics analysis to identify specific mutations in their genomic DNA. The Oxford Nanopore sequencing technology on the MinION device with Field Sequencing Kit SQK-LRK001 and Flow Cell R9.4.1 FLO-MIN106D reagents will be used. The protocol currently used in the project allows optimization of the sequencing process, and the above-mentioned reagents increase the amount of good-quality data provided to the server created as part of the project, thus ensuring its correct configuration and operation, as well as the probability of identifying subsequent gene mutations of the analyzed Bacillus sp. having the influence on the improvement of specific enzymatic activities. The solution used in the project allows to maintain a very good quality of the obtained genomic DNA libraries resulting from the dedicated Oxford Nanopore reagents included in the MinION Starter Packs.
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