hydrophila produce virulence factors such as haemolysin, aerolysin, proteases, lipases
DNAase and enterotoxins, which are important in the pathogenesis of human and fish disease (14, 15, 16).
To perform reverse transcription, RNA was added to the samples of RNA (1000ng/[micro]l) with 0.2 of
DNAase (Invitrogen Life Technologies, Carlsbad, CA, USA) at 37[degrees]C for 5 minutes followed by heating at 65[degrees]C for 10 minutes.
Briefly, lungs were chopped into small pieces with scissors, immersed in 5 mL of collagenase D (200 unit/mL), [Ca.sup.2+] and [Mg.sup.2+],
DNAase, and incubated at 37[degrees]C with shaking for 60 min.
Plasmid DNA in a groundwater aquifer microcosm-adsorption,
DNAase resistance and natural genetic transformation of Bacillus subtilis.
Selected SA cultures were confirmed further, using
DNAase and tube coagulase tests.
Biochemical characterization of the bacterial isolates was done by catalase test, methyl red test, Oxidase test, Indole test, Voges proskauer test,
DNAase test, Sudan III test and Motility test using Sulfide Indole Motility agar (Tittsler and Sandholzer, 1936).
To ensure no DNA contamination, clean-up of the RNA was performed using QIAGEN spin columns with an additional
DNAase step (QIAGEN, Chatsworth, CA, USA).
The
DNAase I treated RNA was again cleaned with the GeneJET RNA Purification kit, re quantified and stored at -80[degrees]C until used.
The Sta-phylococci were further categorized as Staphylococcus aureus and Staphylococcus epidermidis based on Co-agulase and
DNAase tests and sensitivity to Novobio-cin (5 ug) disc.
Total RNA from gill and muscle were extracted using Unizol reagent kit (Biostar, Shanghai, China) according to the protocol, and then the extracted RNA was treated with
DNAase I.