TABLE 1.

Evaluation of 18S rRNA published gene-targeting primer pairs using the SILVA database 18S rRNA genes of rumen protozoa¹.

	RP841F/Reg1302R (Kittelmann et al., 2015)			P-SSU-316F/GIC758R (Ishaq and Wright, 2014)			GIC1080F/GIC1578R (Ishaq and Wright, 2014)			P-SSU-54F/P-SSU-1747R (Sylvester et al., 2004)			P.324f/P.1747r_2 (Zhang et al., 2015)
Genus targeted	Eligible target sequences2	Coverage (%)3	Specificity (%)4	Eligible target sequences	Coverage (%)	Specificity (%)	Eligible target sequences	Coverage (%)	Specificity (%)	Eligible target sequences	Coverage (%)	Specificity (%)	Eligible target sequences	Coverage (%)	Specificity (%)
Charonina	3	0	0	2	50.0	100	3	33.3	99.9	2	100	99.9	3	33.3	100
Dasytricha	22	72.7	99.9	12	100	100	20	75.0	99.9	5	100	99.9	16	37.5	100
Diplodinium	34	100	99.9	17	88.2	100	33	54.5	99.9	15	100	99.9	0	0	NA
Diploplastron	2	100	99.9	1	100	100	2	100	99.9	1	100	99.9	0	0	NA
Enoploplastron	1	100	99.9	1	100	100	1	100	99.9	1	100	99.9	0	0	NA
Entodinium	201	89.1	100	38	92.1	100	196	80.1	99.9	16	100	99.9	175	20.0	100
Eodinium	3	100	99.9	3	100	100	0	0	NA	0	0	NA	0	0	NA
Epidinium	3	100	99.9	2	100	100	3	100	99.9	2	100	100	0	0	NA
Eremoplastron	5	40.0	99.9	5	80.0	100	5	80.0	99.9	4	75.0	99.9	0	0	NA
Eudiplodinium	36	100	99.9	34	100	100	36	33.3	99.9	33	97.0	100	0	0	NA
Isotricha	8	0	0	6	100	100	8	75.0	99.9	6	100	99.9	0	0	NA
Metadinium	6	100	99.9	4	100	100	6	66.7	99.9	4	100	99.9	0	0	NA
Ophryoscolex	43	93.0	99.9	31	96.8	100	43	41.9	99.9	30	100	100	46	2.2	100
Ostracodinium	5	100	99.9	3	100	100	5	100	99.9	3	100	99.9	0	0	NA
Polyplastron	27	88.9	99.9	21	90.5	100	27	22.2	99.9	20	95.0	99.9	30	6.7	100
uncultured	17	94.1	99.9	0	0	NA	17	94.1	99.9	0	0	NA	17	64.7	100
Total	416	88.25	99.9	180	94.4	100	405	66.2	99.9	142	97.9	99.9	287	19.5	100

¹TestPrime 1.0 was used to evaluate the primer pairs (Klindworth et al., 2013).²Eligible target seqs = number of target sequences with corresponding regions of the primer pairs.³Coverage (%) = (number of sequences matched/number of eligible target sequences) × 100. Total coverage mean (bottom row of data) among all genera is weighted for the numbers of sequences within each genus.⁴Specificity (%) = 100 - (outgroup matched sequences/outgroup matchable sequences) × 100. NA = not available because it could be calculated only when at least one sequence was matched. NA values were excluded from the total mean.⁵With one mismatch allowed (and not compromising specificity), coverage of this primer pair equals 95.9%.