Female-specific SNP markers provide insights into a WZ/ZZ sex determination system for mud crabs Scylla paramamosain, S. tranquebarica and S. serrata with a rapid method for genetic sex identification

Table 2 Primers used for the validation of female-specific SNP markers, the extension of female-related sequence and the PCR-based genetic sex identification method

Name	Reference sequence	Primers (5′-3′)	Annealing temperature (°C)
4307 ^a	Cluster_121142	F: TTTGCTTTTTTTGTCTTATGGTTC	52
4307 ^a	Cluster_121142	R: AAACAAATTTACTGAAAACGTGTCT
8966 ^a	Cluster_136335	F: TATAGAGTGCTTTGCATCAATT	51
8966 ^a	Cluster_136335	R: TTCAAAACAAAATTACTGAAAAC
6579 ^a	Cluster_164559	F: CCTTGTGGTTCTTTTGAAC	50
6579 ^a	Cluster_164559	R: AACGAAATTACTGAAAACTTG
9673 ^a	Cluster_22418	F: TCATAGGTACCAAGATGCC	55
9673 ^a	Cluster_22418	R: GGTTTCTCGTAATGTCGTT
8171 ^a	Cluster_31265	F: GAAAACGTGTCTTCCCAGTG	54
8171 ^a	Cluster_31265	R: ATATACACGAAGGTTTGCGTT
11,508 ^a	Cluster_384014	F: GCTTATCATAGTTATTGCCTTGT	53
11,508 ^a	Cluster_384014	R: TGCACTCATGCTGGATTTT
1888 ^a	Cluster_39896	F: GATTTCATCATCACCGACG	57
1888 ^a	Cluster_39896	R: GCAATTCTTGTCTGAGCATG
4225 ^a	Cluster_4545	F: CAGCCCCGACATTAAGGC	56
4225 ^a	Cluster_4545	R: ATATACTGCAATTCTAATGCCAGG
9132 ^a	Cluster_4856	F: ATTATTCTGGTGACTAACA	55
9132 ^a	Cluster_4856	R: GCTAAAACTTCTTTATAGAG
5476 ^a	Cluster_71436	F: CTATATTGTTAATTGTTTTGGTGAC	53
5476 ^a	Cluster_71436	R: TCATCTTCATAGGTACCAATATCA
FLFE-1 ^b	Genome sequence	F: GTACTCTTTAATCAGTTTGTGCCCAT	53
FLFE-1 ^b	Genome sequence	R: CTGCCAGTGATTCAGTGACTTAGC
FLFE-2 ^b	Genome sequence	F: ATGTTTATTGTGTTGTTCAGTGTTGTCT	53
FLFE-2 ^b	Genome sequence	R: CGAGGGTTACTGTAGTTAATGGC
SPC ^c	Extended sequence of female-specific fragment	F: GTTCTGCTTATCATAGTTATTGCCTTG	65
SPC ^c	Extended sequence of female-specific fragment	R: CTGCCAGTGATTCAGTGACTTAGC
SPFS ^c	Cluster_384014	F: CTTAGTATATCACAACTACATCAGGATGT	65
SPFS ^c	Cluster_384014	R: AAGATGCTTGCTGTCTCATTGGT

^aPrimers used for the development and validation of female-related SNP markers; ^b Primers used for the extension of female-related sequence, FLFE: female-related fragment extension; ^c Primers used for PCR-based genetic sex identification method, SPC: Scylla paramamosain control primer, SPFS: S. paramamosain female-specific primer, letters in bold were four mismatch nucleotides

ISSN: 1471-2164