Norgen rna dna purification kit

Swabs from the tarsal conjunctiva were collected at baseline and at one, six- and twelve-months post-surgery. A sterile polyester swab (Puritan) was passed across the conjunctival surface four times, with a quarter-turn on each pass. Swabs were stored in RNAlater (Thermo Fisher Scientific) at 4°C overnight and then long-term at -80°C. Swabs were shipped (on dry ice) to Kilimanjaro Christian Medical Centre (KCMC), Tanzania for processing. RNA and DNA were extracted from swabs using Norgen RNA/DNA purification kit (Norgen Biotek) following manufacturer’s instructions.

At the first time-point, DNA was extracted using the PowerSoil DNA Isolation Kit (Mo Bio Laboratories, USA) from swab samples stored in dry tubes and C. trachomatis was detected by droplet digital PCR, as previously described[5 (link), 9 (link)]. At all subsequent time-points, DNA was extracted from samples stored in RNAlater using the Norgen RNA/DNA purification kit (Norgen Biotek) and C. trachomatis was detected by triplex quantitative PCR (qPCR) for chlamydial chromosomal (omcB) and plasmid (pORF2) genes and a human endogenous control gene (RPP30), as described previously[10 (link)]. Time-point 2 Norgen-extracted samples were tested by both detection methods and the kappa score for agreement was 0.84. Samples were tested in duplicate and were defined as C. trachomatis positive if RPP30 and pORF2 and/or omcB amplified in <40 cycles in one or both replicates.