Analysis and validation of silica-immobilised BST polymerase in loop-mediated isothermal amplification (LAMP) for malaria diagnosis.

ABSTRACT: Bacillus stearothermophilus large fragment (BST_LF) DNA polymerase is reported, isolated on silica via a fused R5 silica-affinity peptide and used in nucleic acid diagnostics. mCherry (mCh), included in the fusion construct, was shown as an efficient fluorescent label to follow the workflow from gene to diagnostic. The R5 immobilisation on silica from cell lysate was consistent with cooperative R5-specific binding of R5₂-mCh-FL-BST_LF or R5₂-mCh-H10-BST_LF fusion proteins followed by non-specific protein binding (including E. coli native proteins). Higher R5-binding could be achieved in the presence of phosphate, but phosphate residue reduced loop-mediated isothermal amplification (LAMP) performance, possibly blocking sites on the BST

SUBMITTER: Seevaratnam D 

PROVIDER: S-EPMC9163865 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature