Han, Sang-Yun
(Department of Marine Biology, Pukyong National University)
,
Rho, Sum
(Haechunma Co., Ltd.)
,
Noh, Gyeong Eon
(Genetics and Breeding Research Center, National Institute of Fisheries Science)
,
Kim, Jin-Koo
(Department of Marine Biology, Pukyong National University)
Interspecific hybridization experiments were conducted between the common seahorse Hippocampus kuda (male) and the slender seahorse H. reidi (female) during artificial rearing to develop a new aquarium fish with unique polyandrous mating. Molecular analysis via mitochondrial DNA (mtDNA) cytochrome b...
Interspecific hybridization experiments were conducted between the common seahorse Hippocampus kuda (male) and the slender seahorse H. reidi (female) during artificial rearing to develop a new aquarium fish with unique polyandrous mating. Molecular analysis via mitochondrial DNA (mtDNA) cytochrome b and nuclear DNA (ncDNA) ribosomal protein S7 gene supported the hybridization between the two species, and the hybrid also showed morphological characteristics of both species. Juveniles of H. kuda have dense melanophores on the whole body or only on the trunk and tail, whereas juveniles of H. reidi have thin melanophores on the whole body or present in stripes only along their prominent trunk and tail rings. However, all the hybrid juveniles had dense melanophores only on the tail, with the striped trunk rings, thus showing an intermediate pattern, and these patterns were limited to the fairly early stage of development (1-10 days old). In contrast, the two eye spines in the hybrid were apparent after 9 days old, which were not inherited from H. kuda (one eye spine), but from H. reidi (two eye spines). According to LOESS (local regression) analysis, the growth rate increased between 20 and 25 days, and the hybrids grew faster than H. kuda when they entered the explosive second phase of growth between 25 and 45 days for all the seahorses. This study highlights the hybridization between H. kuda and H. reidi may contribute to the improved taxonomic information of young seahorses.
Interspecific hybridization experiments were conducted between the common seahorse Hippocampus kuda (male) and the slender seahorse H. reidi (female) during artificial rearing to develop a new aquarium fish with unique polyandrous mating. Molecular analysis via mitochondrial DNA (mtDNA) cytochrome b and nuclear DNA (ncDNA) ribosomal protein S7 gene supported the hybridization between the two species, and the hybrid also showed morphological characteristics of both species. Juveniles of H. kuda have dense melanophores on the whole body or only on the trunk and tail, whereas juveniles of H. reidi have thin melanophores on the whole body or present in stripes only along their prominent trunk and tail rings. However, all the hybrid juveniles had dense melanophores only on the tail, with the striped trunk rings, thus showing an intermediate pattern, and these patterns were limited to the fairly early stage of development (1-10 days old). In contrast, the two eye spines in the hybrid were apparent after 9 days old, which were not inherited from H. kuda (one eye spine), but from H. reidi (two eye spines). According to LOESS (local regression) analysis, the growth rate increased between 20 and 25 days, and the hybrids grew faster than H. kuda when they entered the explosive second phase of growth between 25 and 45 days for all the seahorses. This study highlights the hybridization between H. kuda and H. reidi may contribute to the improved taxonomic information of young seahorses.
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제안 방법
2013). A neighbor-joining tree was constructed from 696 bp of the cytochrome b gene using MEGA, and confidence levels were assessed with 1000 bootstrap replications. Heterozygosity of ncDNA causes a mixed signal as double peaks of sequence chromatograms, and single nucleotide polymorphism (SNP) and insert/deletion (indel) overlap may reveal hybridization (Sousa-Santos et al.
Measurements were made with the microscope-integrated Active Measure software (Shinhanoptics, Seoul, Korea) based on set points for the following parameters: head length (HL), trunk length (TrL), tail length (TaL), snout length (SnL), snout depth (SnD), head depth (HD), dorsal and pectoral fin base lengths (DfL and PfL, respectively), eye diameter (ED), and standard length (SL) (Lourie 2003; Choo and Liew 2006). We derived age–SL relational expression and allometric relational expression (SL–HL, SL–TrL, SL– TaL, SL–SnL, SL–SnD, SL–HD, SL–DfL, SL–PfL, and SL– ED) using LOESS (local regression) curves in ggplot2 package of R software ver.
SYH performed the morphological and molecular experiments and wrote the manuscript. SR and GEN performed the rearing experiment and sampling. SR and JKK suggested all aspects of the study design and commented on the earlier drafts of the manuscript.
SYH performed the morphological and molecular experiments and wrote the manuscript. SR and GEN performed the rearing experiment and sampling.
The PCR was designed to amplify the mtDNA cytochrome b gene using primers Shf2 (5′-TTGCAACCGCATTTTCTTCAG-3′) and Shr2 (5′-CGGAAGGTGAGTCCTCGTTG-3′) under the following conditions: initial denaturation at 94 °C for 2 min 30 s, 35 cycles of denaturation at 94 °C for 30 s, annealing at 50 °C for 30 s, and extension at 72 °C for 75 s, with a final extension at 72 °C for 5 min (Lourie and Vincent 2004), and ncDNA ribosomal protein S7 gene (1st intron, RP1) using primers S7RPEX1F (5′-TGGCCTCTTCCTTGGCCGTC-3′) and S7RPEX2R (5′-AACTCGTCTGGCTTTTCGCC-3′) under the following conditions: initial denaturation at 95 °C for 1 min, 30 cycles of denaturation at 95 °C for 30 s, annealing at 60 °C for 1 min, and extension at 72 °C for 2 min, and a final extension at 72 °C for 10 min (Chow and Hazama 1998).
2016). Therefore, 571 bp of ribosomal protein S7 gene via forward and reverse reading was used for demonstration of hybridization in this study (Fig. 1).
대상 데이터
kuda offspring for morphological analysis. Six specimens were used for a molecular analysis: one each of the H. kuda and H. reidi broodstock, two H. kuda offspring, and two hybrid offspring. We preserved and fixed the specimens in 99% ethanol or 10% formaldehyde.
성능/효과
In contrast, measurements of TrL, TaL, SnD, HD, DfL, and PfL did not differ on LOESS analysis, suggesting that it is difficult to distinguish them based on the allometric patterns in these traits. In two of the three measurements that did change after hybridization (HL, SnL), the curves for allometric growth were higher level for H. kuda than for the hybrid. However, the slope for one of these three measurements (ED) was steeper in the hybrid than in H.
reidi, respectively. Thus, the maternal molecular mtDNA data indicated that the hybrid offspring corresponded to H. reidi and that the eye spine phenotype was inherited from H. reidi, although the hybrid specimens were born from the male H. kuda brood pouch (Fig. 3). Analysis of S7 sequences with a length of 571 bp revealed an average of one indel and 13 SNP overlaps, and these overlaps representing double peaks in the sequence also demonstrated the hybridization of both species (Fig.
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