Gene expression studies for the analysis of domoic acid production in the marine diatom Pseudo-nitzschia multiseries

Table 5 RT-qPCR target gene primer sequences and characteristics

Predicted gene product	Primer sequence	GC (%)	Tm (°C)^a	Amplicon (bp)	Ex-Ex Spanning^b	Efficiency (%)	R²
Cycloisomerase	F: TCATAGGTGGCGTCAAGAACGTGT	50.0	60.3	127	No	99.5	0.995
	R: TCAGCTTGTCGTGCCGAAATTGTG	50.0	60.3
SLC6	F: TCGGACACTACGGAGACTACG	57.1	57.1	73	No	104.2	0.997
	R: ACCAAGGTGAAGGCGACG	61.1	58.0
SLC6 Ex-Ex	F: CATGCACGATACTGTCTATTTCG	43.5	53.6	122	Yes	100.0	0.998
	R: CGTCCAACCAAAATAAGCCAGC	50.0	57.0
Aldo-keto reductase	F: GAATGGGCTACGGAGAGACG	60.0	57.3	114	No	99.5	0.998
	R: GTACAGGCGTGAATTTGGTAGC	50.0	56.2
sHSP	F: GACGAAGGATTCATCACCGTCG	54.5	57.7	141	No	102.9	0.998
	R: GACACCGTTGTCGAGGGTAG	60.0	57.4
PEPCK	F: GCATTGCTCTGCAAACGTCG	55.0	57.7	107	No	100.0	0.997
	R: CAATCAAGGCTCGGTGAGGATC	54.5	57.7
GDH	F: CAATGCCATCAACGCCATCAAGGA	50.0	60.2	128	No	98.4	0.998
	R: CAAAGCCGAGGTTGGCAAGAGTTT	50.0	60.3
PFK	F: CGAGGTGGCATCCAAACGATTGC	56.5	61.1	84	No	105.1	0.998
	R: GCAGCCTGTGTATTGGTATCGTCG	54.1	59.7
PFK Ex-Ex	F: GGAGAAAATCCGCTCGAGGTG	57.1	57.9	111	Yes	99.4	0.997
	R: CTTTGAGAGAACCGCAGCCTG	57.1	58.4
FCP	F: CGTCTCATACCACGGCAC	61.1	55.7	184	No	97.8	0.997
	R: CTTGGATTGATGGTCCACGAG	52.3	55.6

^aThe annealing temperature for all standard curve analyses was performed at 60°C to demonstrate the efficiency of the primer sets under our assay conditions; the calculated Tm values are provided for reference.
^b'Ex-Ex spanning’ refers to primers that span an exon-exon junction; these primer sets did not yield a product using gDNA as a template.

ISSN: 1471-2199