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International Journal of Aquaculture 2012, Vol.2, No.2, 5
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maxima
(11.1%, 2 of 18) (Wang et al., 2009a), and
hard clam
Mercenaria mercenaria
(21.2%, 11/52)
(Wang et al., 2010a). As far as we can determine, high
frequency of duplicated locus linked in tandem such
as the 30.8% observed in this study has not been
reported in any mollusk. This finding implies that the
complex duplications of
B. areolata
genome are
partly characterized by overlapping and/or linkage
in tandem of microsatellites, mini-satellites and
satellites.
Table 2 Information of nine
Babylonia areolata
duplicated loci
Locus
GenBank acc. no.
Repeat motif
§
Primer sequences (5'
-
3')
Ta(
℃
) MgCl
2
(mM) as
n
F: taggtgcaggccatatcagag
HUBA02*
FJ594999
(AC)
N
(CT)
N
R: gtgaaacggtcaaatgttcaag
62
1.5
275-444 32
F: gatcacgatttgaaagactgaaag
HUBA04
FJ595001
(GT)
N
R: cggaacttgcaacatttaacac
60
1.5
191-311 32
F: aagctcgttcaacttcacgtc
HUBA05
FJ595002
(GT)
n
(GA)
N
R: gatcaatccgctgtctttctg
62
1.5
271-451 32
F: atcgccacaatgacaatcaac
HUBA06*
FJ595003
(CA)
N
(TCA)
N
F: actggatgatgcgatgacaac
50
1.5
404-551 32
F: gaaacttcaaacgacaccaaac
HUBA11*
FJ595008
(GT)
N
(CT)
n
R: gaaagagacgaagggtggaag
61
1.5
286-414 32
F: agtacgggtttcttctttgctc
HUBA12
FJ595009
(GACA)
N
(GA
n
R: agcctgcgaatctgattactg
60
1.5
277-425 30
F: tgccaaatttgttgtcaactg
HUBA14
FJ595011
(AC)
N
R: cttccgtcaacagtgaccttg
60
1.5
195-285 31
F: tgcctaatttggtgtcaactg
HUBA16*
FJ595013
(AC)
N
R: cttgctttcggtcactttttg
60
2.0
166-278 29
F: ttttctgcacatgaagaaccac
HUBA19
FJ595016
(TG)
n
R: aaaaacctccaccctttaccc
53
2.0
242-508 32
Note: The number of individuals analyzed (n), allele size across multi-locus range in base pairs (as) are presented.
§
Repeat motif: N,
pure; n, interrupted;
*
duplicated loci amplified parallel loci clearly linked in tandem
There probably is high frequency of microsatellite
sequences in
B. areolata
genomes, since we obtained
an enrichment efficiency of 97.5% (118 of 121) in this
study, which is much higher than that obtained in
other four molluscan species whose microsatellite
markers were developed in parallel in our laboratory
using the same protocol and reagents: 63.3% for
Spisula solidissima
(Wang et al., 2009b)
,
53.2% for
Pinctada maxima
(Wang et al., 2009a), 59.4% for
Mercenaria mercenaria
(Wang et al., 2010a), and
58.5% for
Haliotis diversicolor
(Wang et al., 2011c).
Extraordinarily, there are sixty-two microsatellite
sequences (52.5%, 62 out of 118) contained too long
and complex microsatellites to have sufficient
flanking regions for primer designing, such high
frequency was also reported in
B. areolata
(Chen et al.,
2009; Wang et al., 2011a) and other whelk e.g.
Buccinum undatum
(Weetman et al., 2005). These
results suggested that the
B. areolata
genome may be
a good model to study the evolution of genome
duplication in mollusk.
The 4 polymorphic microsatellite markers developed
here are useful addition to the collection of other
molecular markers that are now available for
B.
areolata
, including mitochondrial markers and other
microsatellites markers (Chen et al., 2009; Wang et al.,
2011a). They will prove valuable for future population
genetic studies and in tracking of hatchery strains in
this species.
The finding of significant proportions of locus
duplication suggests that the use and isolation of
microsatellite markers in
B. areolata
may be
challenging. Perhaps further efforts in this species
should be directed to the development of SNP markers
based on the second generation sequencing technology.