International Journal of Aquaculture, 2013, Vol.3, No.19, 105
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advantageous as a preliminary tool for studying the
population structure of penaeids with microsatellite
analysis. RAPD technique designed to detect sequence
changes within the priming site as well as differences
in length, through insertion or deletion, between them.
Mutation at the priming site will affect primer
annealing and amplification of particular segment
(
Caetano-Anolles and Bassam, 1993), whereas deletion
or insertion between primering site will alter the
length of amplified fragment, both instances will
result in changes in the RAPD profile produced by a
given primer, and significance of these differences can
be used to determine the extent of population structure.
RAPD fragment represent a combination of product
amplified from coding and non-coding loci, estimator
of genetic diversity extracted from them tend to be
more informative of the overall genetic variation in
the species than those derived from coding loci alone.
The population separated by thousands of kilometers
can be genetically more similar than others over very
short distances and differences between species in
level of genetic variation, and genotype distribution
might be related to life history types (Mulley and
Latter, 1980; De La Rosa-Velea et al., 2000).
Previously many genetic diversity studies using
allozyme analysis has been used to determine the
levels of variation and degree of genetic subdivision
for several shrimp species (Lester and Pante, 1992; De
la Rosa-Velez et al., 2000; Gracia-Machado et al., 2001;
Gusmao et al., 2005), and mitochondrial DNA
restriction fragment length polymorphism (Benzie et
al., 1993; Klinbunga et al., 1999, 2001; Gracia-Machado
et al., 2001). Genetic diversity and geographic
differentiation of the
P. monodon
mitochondrial COI
sequences (Kumar et al., 2007; Khamnamtong et al., 2009).
Randomly amplified polymorphic DNAs (Aubert and
Lithtner, 2000; Mishra et al., 2009; Lakra, 2010;
Gilkolaei et al., 2011) and microsatellite (Wolfus et al., 1997;
Brooker et al., 2000; Supungal et al., 2000; Maggioni
et al., 2003; Robainas-Barcia and García-Machado, 2012;
Mandal et al., 2012). In all studies, the major penaeid
were
Litopenaeus vannamei
,
P. monodon
,
and
Masupenaeus japonicus
,
but in the present study, the
most available and economically important penaeid
populations were the
candidate species.
Previously
no report was available on the molecular characterization
of
Fenneropenaeus indicus
,
Penaeus semisulcatus
,
Penaeus monodon
of the south east coast region of
India, and hence an attempt has been made to reveal
the genetic diversity of three penaeid population in
south east coast of India.
1
Results
Among 15 numbers of primers used for this study, 3, 4,
and 6 number of primers, worked well for
F. indicus
,
P.
semisulcatus
and
P. monodon
respectively. The primer
sequences for this study (Table 1). Among the primer
RM 03, 04, 05, 07, 11, 14, 15, 17 used for present
study the RM 03, 04, 07 primers amplified the DNA
product of three species taken for genetic analysis.
DNA bands size produced 1000 to 10000 bp. For this
1000
Kbp maker were used as reference and
polymorphic profile for
F. indicus
,
P. semisulcatus
,
and
P. monodon
shown as Figure 1, Figure 2, and
Figure 3 respectively. The total number of bands,
number of polymorphic bands, number of monomorphic
bands and percentage of polymorphic bands for all
species collected from all location (Table 2) and genetic
diversity value (Table 3) were counted.
Table 1 Primer used for present study
S.No
Primer name
Sequence
1
RM03
5’
AAT CGG GCT G 3’
2
RM04
5’
GAAACG GGT G 3’
3
RM05
5’
GTG ACG TAG G 3’
4
RM07
5’
CAA TCG CCG T 3’
5
RM011
5’
TCC TCAAGA C 3’
6
RM014
5’
GTA TTG CCC T 3’
7
RM015
5’
TTC CCC GAC C 3’
8
RM017
5’
TCC CTC GTG C 3’
Figure 1 RAPD profile for
F. indicus
Figure 2 RAPD profile for
P. semisulcatus