Genetic enhancement of Bacillus cyclodextrin glycosyltransferase production

The bacterial isolates obtained from the botanical garden soil of the National Research Center in Dokki, Giza, Egypt, showed high levels of cyclodextrin glycosyltransferase (CGTase) activity. The CGTase superior Bacillus (WT3) was classified as Bacillus hunanensis using the 16S-ribosomal RNA sequence and the Basic Local Alignment Search Tool (BLAST). The mutagen ethyl methane sulfonate (EMS) was used to create CGTase mutants of WT3 that produced more. After the testing of 30 mutants chosen in a screening assay, the best mutant for CGTase biosynthesis was WT3-60-8, which produced 84.19 U/mL with a 110.6% relationship to the parental-strain. Protoplast fusions between WT3-60-4 and WT3-60-8 mutants were carried out to increase CGTase activity. Twenty-seven fusants were collected, with fusant (F1-8) showing a 122.4 percent improvement in CGTase activity over its parental strain. Many different DNA banding patterns were observed while fingerprinting with random amplified polymorphic DNA (RAPD). Furthermore, the genetic backgrounds of the two excellent mutants and four superior fusants, as well as the parental-strain, were divided into three clusters, the phylogenetic tree was obtained, and the genetic backgrounds of the two excellent mutants and four superior fusants, as well as the parental-strain, were detected based on genetic distances.

Automated summary

Bacterial isolates from the soil of the botanical garden in Egypt showed high levels of CGTase activity. The superior Bacillus strain WT3 was classified as Bacillus hunanensis. Mutants of WT3 were created using EMS, and the best mutant for CGTase biosynthesis was WT3-60-8. Protoplast fusions between mutants WT3-60-4 and WT3-60-8 resulted in fusant F1-8 with significantly improved CGTase activity. DNA fingerprints using RAPD showed different banding patterns, and the genetic backgrounds of the mutants and fusants were analyzed based on genetic distances.