Development of a versatile TaqMan™ real-time quantitative PCR (RT-qPCR) compliant anchor sequence to quantify bacterial gene transcripts from RNA samples containing carryover genomic DNA

Table 3 Quantification of RNA extracted from soil spiked with two concentrations of Pseudomonas sp. LBUM300 after multiple rounds of DNase I treatment

Days post inoculation	Bacterial spiking amount	RNA amount post DNase-I treatment (ng/μL)^*¶
Days post inoculation	Bacterial spiking amount	Round 0	Round 1	Round 2	Round 3
Day 0	Control	33.7 ± 1.7	28.2 ± 2.4	25.2 ± 1.7	16.5 ± 2.2
	Control	33.7 ± 1.7	(-16.3%)	(-25.2%)	(-51.0%)
	10⁷ bacteria/mL	53.9 ± 2.4	45.3 ± 3.1	40.1 ± 3.8	34.7 ± 4.2
	10⁷ bacteria/mL	53.9 ± 2.4	(-15.9%)	(-25.6%)	(-35.6%)
	10⁹ bacteria/mL	102.4 ± 2.9	82.2 ± 4.4	61.7 ± 3.3	40.7 ± 4.9
	10⁹ bacteria/mL	102.4 ± 2.9	(-19.7%)	(-39.7%)	(-60.2%)
Day 7	Control	34.4 ± 2.3	30.7 ± 2.8	26.8 ± 3.8	22.2 ± 4.5
	Control	34.4 ± 2.3	(-10.7%)	(-22.1%)	(-54.9%)
	10⁷ bacteria/mL	56.6 ± 3.2	43.8 ± 4.4	32.7 ± 4.6	29.1 ± 3.5
	10⁷ bacteria/mL	56.6 ± 3.2	(-22.6%)	(-42.2%)	(-48.5%)
	10⁹ bacteria/mL	59.5 ± 2.5	42.7 ± 3.2	34.9 ± 4.7	23.1 ± 5.4
	10⁹ bacteria/mL	59.5 ± 2.5	(-28.2%)	(-41.3%)	(-61.1%)

* : Avg of 3 replicate samples ± standard error of mean.
^¶: values in brackets indicate % reduction in RNA amounts (with respect to original starting amount), recovered after each round of DNase-I treatment.

ISSN: 1472-6750