Identification of candidate single nucleotide polymorphic markers in brown-marbled grouper Epinephelus fuscoguttatus (Forsskal, 1775)
Brown -marbled grouper Epinephelus fuscoguttatus is a premium marine food fish with high demand in Asia. However, its aquaculture development is limited by broodstock availability with heavy reliance on wildstocks. The growth and stress polymorphic markers were explored using high-throughput sequ...
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| Format: | Thesis |
| Language: | English |
| Published: |
2021
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| Subjects: | |
| Online Access: | http://psasir.upm.edu.my/id/eprint/99225/1/IB%202021%201%20IR.pdf http://psasir.upm.edu.my/id/eprint/99225/ |
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| Institution: | Universiti Putra Malaysia |
| Language: | English |
| Summary: | Brown -marbled grouper Epinephelus fuscoguttatus is a premium marine food fish
with high demand in Asia. However, its aquaculture development is limited by
broodstock availability with heavy reliance on wildstocks. The growth and stress
polymorphic markers were explored using high-throughput sequencing and the
genetic diversity of E. fuscoguttatus for potential use in selective breeding to
facilitate the development of aquaculture. Thus, the first objective of this study was
to investigate the salinity tolerance of E. fuscoguttatus and its effects on the serum
cortisol levels, survival and growth in an aquaria setting. This was a preliminary
study towards the identification of SNP markers later on. In this study, grouper
juveniles (92.43 ± SEM 0.51 mm) were maintained in 31 ppt seawater for one-week
acclimatization and transferred into five tanks with different seawater dilutions (5,
10, 15, 20, 25 ppt). The results revealed that serum cortisol of fish in high change of
salinity (from 31 ppt to 15, 10 and 5 ppt) was significantly higher than the control
group immediately after exposure. In the higher salinity change (from 31 to 5 ppt),
the survival percentage was 50%, while no mortality observed in the lower change
of salinity. Whereas after 2 weeks, a significant difference in the decrease of mean
weight of fish in the higher change of salinities (from 31 to 15 ppt, 10 ppt, and 5 ppt)
was observed compared to the control group indicating the effects of chronic stress
on the growth performance of fish. In the aquaria experiment, some fish were more
stress-tolerant than others could be used as potential as candidate for selective
breeding. The second objective of this study was to measure the growth performance
of E. fuscoguttatus in sea cage culture to detect the slow and fast growers for DNA
sequencing. The fish were grown for 10 months in a floating cage where the
morphometrics of 100 fish (215.10 ± SEM 21.64 mm) were measured and 20 fish
were intramuscularly microchipped. At the end of 10 months, 10 fish recovered with
microchip were categorized based on their size as fast- and slow-growers and
processed for DNA extraction. The next objective was to assess the potential of a novel high-throughput sequencing method on the muscle DNA samples. Highthroughput
sequencing using the double digestion restriction associated DNA
sequencing (ddRADSeq) was done utilising two restrictive enzymes: PstI and MspI.
The results showed 146,244 sequences and with 16172 SNP of 25 or less per
sequence. Out of the 16172 SNP, 64% were identified with one SNP, while 5776
(35%) were identified with more than two SNPs. An overall FST for the two
populations of fast- and slow-growers was extremely low (0.0705) with 21.2% -
26.1% polymorphic loci and high observed homozygosity as calculated by STACKS
indicating inbreeding. The last objective of this study was to analyse the genetic
diversity of E. fuscoguttatus and isolate and identify SNP markers using
bioinformatic tools as candidate markers. Analysis of the 382 filtered SNP sequences
using BLAST, GO and UniProt databases revealed 36 sequences which matched the
Epinephelus spp. and 18 GO biological processes of 57 genes in the fast-and slowgrowers
related to growth and stress. Sequencing using the novel ddRAD Seq
revealed four genes: acss2, abhd15, lrp2 and pdp1 as potential candidate markers
that could assist in the selection of broodstock fish with superior traits. This is the
first ddRADSeq performed in this species. The outcome of this study can contribute
to understanding suitable bioinformatics pipelines, further adding to the limited
genetic information available for establishing and identifying suitable markers for
the future of assisted selective breeding of this species. |
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