Aquaculture Research

Research Article

Efficient Production of Doubled Haploids as Isogenic Line Founders via Double Heat Shock Treatment in Willow Gudgeon (Gnathopogon caerulescens)

Table 2

Homo/heterozygosity determination by Gras-Di analysis in some individuals whose zygosity was tentatively determined by STR analysis.


ID	Exp. grp	Ploidy	Putative jdg from 8 MS^†	Gras-Di analysis					Final jdg
ID	Exp. grp	Ploidy	Putative jdg from 8 MS^†	Index sequence^‡	^#seq pair reads	Mean coverage	Hetero (%)^§	Score^¶	Final jdg

#1 Ctrl-01	IC	2n	—	TGTCGAACAA	1,025,334	22.6×	22.2	Diploid	Diploid
#1 Ctrl-02	IC	2n	—	GCGAGGATAA	728,440	20.3×	20.4	Diploid	Diploid
#1 Ctrl-03	IC	2n	—	ACTTCTGGTG	801,061	20.8×	20.4	Diploid	Diploid
#1 G-haploid_01	GC	n	Haploid	CAGGACCTGA	973,994	21.8×	4.7	Haploid	Haploid
#1 G-haploid_02	GC	n	Haploid	ACTCTCACCA	721,766	19.5×	4.9	Haploid	Haploid
#1 G-haploid_03	GC	n	Haploid	GCGACTGTGT	833,217	20.5×	4.8	Haploid	Haploid
#1 Meio-G 2n_01	F25S41	2n	Meio-G 2n	GCCGTGCATA	281,957	16.2×	6.1	Meio-G 2n	Meio-G 2n
#1 Meio-G 2n_02	F25S41	2n	Meio-G 2n	ATCGCCTATA	657,996	16.3×	5.8	Meio-G 2n	Meio-G 2n
#1 Meio-G 2n_03	F29S45	2n	Meio-G 2n	GACACTCCTA	1,236,210	25.1×	4.4	DH	Meio-G 2n
#1 DH_01	F23S39	2n	DH	TAGGTTGGAA	911,283	22.1×	4.6	DH	DH
#1 DH_02	F23S39	2n	DH	ACAATTACGG	868,655	21.0×	4.8	DH	DH
#1 DH_03	F23S39	2n	DH	ATTGTACCGG	990,378	22.5×	4.8	DH	DH
#1 DH_04	F25S41	2n	DH	TAGTGGTCGG	839,948	21.4×	4.8	DH	DH
#1 DH_05	F25S41	2n	DH	CAGAAGATGG	1,019,925	21.8×	4.7	DH	DH
#1 DH_06	F25S41	2n	DH	CGTTATCACA	973,496	21.5×	5.9	Meio-G 2n	Meio-G 2n
#1 DH_07	F27S43	2n	DH	TAATGGTGTG	942,037	18.5×	5.3	Meio-G 2n	Meio-G 2n
#1 DH_08	F27S43	2n	DH	CTGCTAGCTG	916,433	23.9×	4.7	DH	DH
#1 DH_09	F27S43	2n	DH	ACACGTAGCC	985,624	23.4×	4.9	DH	DH
#1 DH_10	F29S45	2n	DH	GAGACCATCC	1,094,788	23.4×	5.2	Meio-G 2n	Meio-G 2n
#1 DH_11	F29S45	2n	DH	CGTGCGATTC	968,233	22.7×	4.8	DH	DH
#1 DH_12	F29S45	2n	DH	GTAGTGGCTT	939,832	22.5×	4.9	DH	DH
#1 DH_13	F31S47	2n	DH	TCGCGCGTAA	821,055	21.3×	5.1	Meio-G 2n	Meio-G 2n
#1 DH_14	F31S47	2n	DH	GGTTCCTATT	917,613	21.6×	4.9	DH	DH
#1 DH_15	F31S47	2n	DH	TACCAAGTGA	1,051,123	23.4×	5.0	Meio-G 2n	Meio-G 2n
Dam#03	—	2n	—	CGCAAGATTA	1,197,446	26.3×	21.5	Diploid	Diploid
#3 Ctrl-01	IC	2n	—	CCTGACAGAG	739,817	22.8×	20.4	Diploid	Diploid
#3 Ctrl-02	IC	2n	—	ACAACCGTCG	958,971	24.8×	20.9	Diploid	Diploid
#3 Ctrl-03	IC	2n	—	ACGCGTTGTG	498,076	20.8×	21.4	Diploid	Diploid
#3 G-haploid_01	GC	n	Haploid	CACATAGTAC	1,063,823	24.3×	4.3	Haploid	Haploid
#3 G-haploid_02	GC	n	Haploid	GTCGATGTCC	819,819	22.6×	4.2	Haploid	Haploid
#3 G-haploid_03	GC	n	Haploid	TAGGCTTCTC	960,208	22.9×	4.2	Haploid	Haploid
#3 Meio-G 2n_01	GC	2n	Meio-G 2n	ACGGTACGTT	862,012	22.8×	17.4	Meio-G 2n	Meio-G 2n
#3 Meio-G 2n_02	F27–	2n	Meio-G 2n	CGACGTTAGA	1,466,196	27.9×	18.9	Meio-G 2n	Meio-G 2n
#3 Meio-G 2n_05	F27S43	2n	Meio-G 2n	CCATAGAGAA	1,552,566	26.8×	4.1	DH	Meio-G 2n
#3 DH_01	F23S39	2n	DH	TGTCTCTCAA	1,680,589	27.9×	4.2	DH	DH
#3 DH_02	F23S39	2n	DH	GCCAAGGAGT	1,160,228	23.9×	4.4	DH	DH
#3 DH_03	F23S39	2n	DH	GTTGTCAGAA	1,286,799	25.1×	4.1	DH	DH
#3 DH_04	F23S39	2n	DH	AACGCTCCGA	1,630,828	26.7×	4.2	DH	DH
#3 DH_05	F23S39	2n	DH	TTCATGGTCA	1,390,779	25.0×	4.2	DH	DH
#3 DH_06	F25S41	2n	DH	GGATTACTCA	1,517,992	24.6×	4.4	DH	DH
#3 DH_07	F25S41	2n	DH	CCACAACATG	1,705,734	28.9×	4.1	DH	DH
#3 DH_08	F25S41	2n	DH	TAGTGCGCTG	973,533	22.6×	4.6	DH	DH
#3 DH_09	F25S41	2n	DH	TGCATGTTAC	1,755,285	27.7×	4.2	DH	DH
#3 DH_10	F25S41	2n	DH	GAACTATTCC	1,357,694	24.8×	4.3	DH	DH
#3 DH_11	F27S43	2n	DH	GTGGACCTTC	1,276,363	24.5×	4.3	DH	DH
#3 DH_12	F27S43	2n	DH	TGTCGGCGAT	984,132	21.9×	4.6	DH	DH
#3 DH_13	F27S43	2n	DH	GAGTACCTGT	1,010,860	22.8×	5.2	Meio-G 2n	Meio-G 2n
#3 DH_14	F27S43	2n	DH	CACAGGTGAA	1,484,192	24.9×	4.4	DH	DH

, ID, individuals for which sufficient quantities of DNA were available were selected for analysis. ^†, putative jdg from 8 MS, estimation by zygosity of eight microsatellite loci. ^‡ index sequence, sequence of reverse primer using as index sequence. ^§ hetero (%), percentage of the number of heterozygous loci out of the number of loci detected. ^¶ score, the hetero (%) of haploids in the Gras-Di analysis results was used as a reference for this determination. final jdg, judgement by both microsatellite loci analysis and Gras-Di analysis. Ctrl, intact control (IC); haploid, haploid induced in gynogenetic control (GC); Meio-G 2n, gynogenetic diploid induced by retention of the second polar body; DH, doubled haploid.