Isolation of oral bacterial DNA for 16S rRNA sequencing to determine nitrate-sensitive oral microbiota following acute beetroot juice supplementation

Dietary nitrate (NO3-) supplementation, provided as beetroot juice, improves cardiovascular, cognitive, and contractile function by augmenting nitric oxide (NO) biosynthesis; however, the specific taxa responsible, and whether NO3- ingestion can promote a favorable microbial composition to produce NO is yet to be established. To determine whether acute NO3- supplementation is associated with specific microbial taxa in females, 15 healthy adult females consumed an acute dose of NO3--depleted beetroot juice (PL) and NO3--rich beetroot juice (BR). Bacterial DNA from the buccal cells were isolated for subsequent 16s DNA sequencing for preserved regions in their 16s rRNA gene. Concentrations were assessed with Nanodrop and Qubit methods, seeking \u3e 20 ng/uL and A260/A280 values of 1.8 to 2.2, in the former. The Nanodrop showed DNA concentrations: (Condition A: 37.66 ± 18.92; vs Condition B: 37.52 ± 11.54 ng/uL) and adequate DNA purity (Condition A: 2.03 ± 0.28; vs Condition B: 2.06 ± 0.21). The Qubit analysis lower than values considered adequate (Condition A: 14.05 ± 9.59; vs Condition B: 13.33 ± 7.22 ng/uL); however, the next step is to amplify the isolated DNA using the PCR method, so the concentration is more of a concern if concentrations remain low following PCR. Upon completion of data collection and isolation for n=40, DNA will be amplified and sequenced to determine whether specific microbial taxa are associated with NO3- ingestion and elevation in plasma NO2- concentrations in blood