Molecular characterization of cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) associated with the erythrocyte antigens in dogs

Uno, Yumiko; Kawakami, Shota; Ochiai, Kazuhiko; Omi, Toshinori

doi:10.1186/s40575-019-0076-1

Canine Medicine and Genetics

Table 3 List of primer used for analysis of the canine CMAH gene

From: Molecular characterization of cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) associated with the erythrocyte antigens in dogs

Amplicon	Primer name	Primer sequences (5′-3′)	Location*	Annealing (°C/s)	Predicted product size (bp)	Genbank Accession No., [Ref]
cDNA cloning
Fragment 1	dCMAH-cE1aF	CTGTTTTGTGCAGTTTGGCCTCTT	exon 1 (5′-UTR)	*55/60	1146	[30]
Fragment 1	dCMAH-cE9R	TTGTCTGATGGATGGGATTCCACA	exon 9			NC_006617.3
Fragment 2	dCMAH-cE8F	CCTGAGAAGGTTGCTCTAATGA	exon 8	*55/60	925	NC_006617.3
Fragment 2	cDMAH-cE15R	TGATCAAGATGTAGCGTCAGTAAAT	exon 15 (3′-UTR)			NC_006617.3
Fragment 3	dCMAH-cE1aF	CTGTTTTGTGCAGTTTGGCCTCTT	exon 1 (5′-UTR)	*55/180	1852	[30]
Fragment 3	cDMAH-cE15R	TGATCAAGATGTAGCGTCAGTAAAT	exon 15 (3′-UTR)			NC_006617.3
SNP discovery
Fragment 1	dCMAH-gE1F	CTCCAGGCTGCCGTCCTTCTA	Promoter	*55/30	227	NC_006617.3
Fragment 1	dCMAH-gE1R	CAACAGCTTTCAGTTCTTGAAT	intron 1			NC_006617.3
Fragment 2	dCMAH-gE2F	GCCTGGATACTTGGAGGGAGG	intron 1	*55/30	393	NC_006617.3
Fragment 2	dCMAH-gE2R	TGATCAGAGAGATGTCCTAAGT	intron 2			NC_006617.3
Fragment 3	dCMAH-gE3F	AGTATTATCTCCTAATGGTTT	intron 2	*55/30	369	NC_006617.3
Fragment 3	dCMAH-gE3R	AAGTCCGACTCTACACAGTTT	intron 3			NC_006617.3
Fragment 4	dCMAH-gE4F	TGAGTTGGTGTTGGTCTTAAG	intron 3	*55/30	483	NC_006617.3
Fragment 4	dCMAH-gE4R	TCCTAAGAAATTTGCTTAATAG	intron 4			NC_006617.3
Fragment	dCMAH-gE5F	TTGAGCATTCTTAGAAGCGAA	intron 4	*55/30	500	NC_006617.3
Fragment	dCMAH-gE5R	CCTCACTCTGAGTGGTATATA	intron 5			NC_006617.3
Fragment 6	dCMAH-gE6F	AATTCCTTGCTTCTTGATCAACA	intron 5	*55/30	334	NC_006617.3
Fragment 6	dCMAH-gE6R	ATCTTAAAGTAACTACCTCATCTAC	intron 6			NC_006617.3
Fragment 7	dCMAH-gE7F	GAAGGATTTCTTTCCAGATGAGC	intron 6	*55/30	254	NC_006617.3
Fragment 7	dCMAH-gE7R	CAGATGGCTGTTATCACCCTT	intron 7			NC_006617.3
Fragment 8	dCMAH-gE8F	GGACATAAGTGATGCTTCTCTA	intron 7	*55/30	445	NC_006617.3
Fragment 8	dCMAH-gE8R	GGCGCTAAACCACTAAGCCAC	intron 8			NC_006617.3
Fragment 9	dCMAH-gE9F	TTCCTGTGTTAGTCTATCCAT	intron 8	*55/30	386	NC_006617.3
Fragment 9	dCMAH-gE9R	AAGCTCCGTGCTCTCAGGCAG	intron 9			NC_006617.3
Fragment 10	dCMAH-gE10F	TAAGTGGATAGGATTGTGAAG	intron 9	*55/30	373	NC_006617.3
Fragment 10	dCMAH-gE10R	CCTACAAATAAGGACTTGTCA	intron 10			NC_006617.3
Fragment 11	dCMAH-gE11F	GATAAGAGAACTTTCCTGTAT	intron 10	*55/30	433	NC_006617.3
Fragment 11	dCMAH-gE11R	TCCGTGAAGCAGTTGGTAGCG	intron 11			NC_006617.3
Fragment 12	dCMAH-gE12F	CATTGCTATCAATTAAGGCTG	intron 11	*55/30	360	NC_006617.3
Fragment 12	dCMAH-gE12R	GTAAAGTAAGGTGTTAACAAT	intron 12			NC_006617.3
Fragment 13	dCMAH-gE13F	AGCCTGTCATATCTACTCCAT	intron 12	*55/30	393	NC_006617.3
Fragment 13	dCMAH-gE13R	GAAATGAAGCCCTAATGGTGG	intron 13			NC_006617.3
Fragment 14	dCMAH-gE14F	CAGTATGGAAGCACCATCTCT	intron 13	*55/30	353	NC_006617.3
Fragment 14	dCMAH-gE14R	ATGTTCTTGCAATGTTAGCTC	intron 14			NC_006617.3
mRNA expression
CMAH	dCMAH-cE2F	TGTGCAAGCATCAAGGAGGC	exon 2	*55/30	342	NC_006617.3
CMAH	dCMAH-cE4R	AGGCAGGACCTATTAACCAG	exon 4			NC_006617.3
GAPDH	dGAPDH-cE1/2	CACAGTCAAGGCTGAGAACG	exon 1/2	*55/30	101	[37]
GAPDH	dGAPDH-cE2	CACCAGCATCACCCCATTT	exon 2			[37]

Number of exon is detemined by comaparison between Ac. No. LC382414 and NC_006617.3 in this study
NC_006617.3: Canis lupus familiaris breed boxer chromosome 35, CanFam3.1, whole genome shotgun sequence
d dog, E: exon, F Forward primer, R reverse primer, number exon number, c designed, g designed from genomic sequence in primer name

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ISSN: 2662-9380

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