Deletion of HindIIR and HindIIIR improves DNA transfer via electroporation toHaemophilus influenzaeRd

ABSTRACT Haemophilus influenzae is a bacterial species of interest for its medical relevance and utility as a model system. Despite its role in several landmark molecular and synthetic biology studies, H. influenzae remains underexplored as a potential chassis organism. The limited availability of reliable and convenient transformation methods and genetic tools for H. influenzae are obstacles to this end. However, a strain of H. influenzae Rd KW20 lacking the type II restriction endonucleases HindII and HindIII has previously been developed. Here, we show that this strain is more readily transformable by electroporation than wild-type Rd KW20. We also developed a series of multi-host plasmids carrying antibiotic selection and fluorescent visual markers based on the pSU20 vector. The availability of H. influenzae ΔHindII/III, paired with the electroporation method and plasmids presented here, will promote the exploration of H. influenzae as a host organism for synthetic biology applications.