Optimizing locked nucleic acid/2'-O-methyl-RNA fluorescence in situ hybridization (LNA/2'OMe-FISH) procedure for bacterial detection.

ABSTRACT: Despite the successful application of LNA/2'OMe-FISH procedures for bacteria detection, there is a lack of knowledge on the properties that affect hybridization. Such information is crucial for the rational design of protocols. Hence, this work aimed to evaluate the effect of three essential factors on the LNA/2'OMe hybridization step-hybridization temperature, NaCl concentration and type and concentration of denaturant (formamide, ethylene carbonate and urea). This optimization was performed for 3 Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa and Citrobacter freundii) and 2 Gram-positive bacteria (Enterococcus faecalis and Staphylococcus epidermidis), employing the response surface methodology and a Eubacteria probe. In general, it was observed that a high NaCl concentr

SUBMITTER: Azevedo AS 

PROVIDER: S-EPMC6544301 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature