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International Journal of Aquaculture, 2013, Vol.3, No.24, 138
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variety of markers for the development of authentic
genetic relationships among closely related species.
The RAPD technique provides an efficient, simple and
inexpensive method of generating molecular data.
Further, it is highly polymorphic marker and does not
require any prior knowledge of the genetic makeup of
the organism (Hadrys et al., 1992). In this study
suitability and reliability of RAPD markers was
assessed for understanding the Phylogenetic
relationships among and within the species of cyprinid
species. In the present study of 12 decamer primers
used to screen DNA samples, 8 (66%) detected
scorable polymorphism in banding pattern among all
the 75 individuals. Eight selected primers generated a
total of 3371 bands of which 3008 were polymorphic.
An example of the representative polymorphic profiles
of 75 individuals with eight primers is shown in
Figure 1. The number of bands per individuals ranged
from 4 to 12 and bands amplified ranged in size from
100-1400 bp. The average number of bands per primer
ranged between 182 (S-131) and 497 (S-159) with a
mean of 421.3. The proportion of polymorphic
markers across the primers ranged between 79.75%
and 97.96% with an average of 89.23% shown.
Rahman et al. (2009) studied genetic variations of
wild and hatchery populations of
Catla catla
by
RAPD markers and found overall 54.55%
polymorphism. Garg et al
.
(2010) have also reported
an analysis for RAPD to assess the extent of genetic
diversity within and between three populations of the
catfish,
Clarias batrachus
and obtained 72 scorable
DNA fragments out of which 68 (86.66%) were
polymorphic. We found that 89.23% of the loci in our
study were polymorphic as compared to the 75%
reported by Islam et al
(2005) in
Catla catla
, 55.76%
in
Oreochromis niloticus.
Zaeem and Ahmed (2006),
64.98% in
Mystus vittatus
by Garg et al. (2009) and
86.66% by Garg et al. (2010) in assessment of genetic
diversity of
Clarias batrachus.
As reported in this study, after a screen of eight
primers, 91.04, 79.7, 95.05, 90.04, 91.14, 97.96, 85.81
and 86.6% polymorphic DNA markers were obtained
for
C.c. communis
,
C.c. specularis and C.carassius
using primers S-71, S-111, S-131, S-145, S-159,
S-161, S-177 and S-187 respectively. The results thus
showed usefulness of RAPD markers for studying
genetic variation in cyprinid
fishes. Species-specific
RAPD profiles between the species under study were
observed using eight random primers. Similar results
were also reported by El-Alfy et al., (2009) assessed
genetic variation among nile tilapine fishes by RAPD
markers. The UPGMA dendrogram obtained from the
RAPD data clearly depicts the relationships among
these species. The highest interspecies genetic
similarity was exhibited between
C.c. communis
and
C.c. specularis
and
supports the hypothesis that these
two cyprinids are closely related. Similar type of study
was also done in Indian major carps PCR-RFLP of
cytochrome b gene has been employed efficiently for
the study of genetic variations, identification and
resolving taxonomical ambiguity of closed related fish
species (Barman et al., 2003).
In our study,
comparable levels of genetic polymorphism
(50.0%, 72.7%, 87.5% and 87.5%) were obtained by
using four restriction endonucleses (MboI, Hin6I,
BsuRI and MspI). Some restriction enzymes like
MboI and BsuRI generated some unique restriction
fragments for the two species, thus indicating the wide
genetic base of the Cyprinid species. The presence of
the unique composite restriction fragments among the
two species indicates the usefulness of the approach
for fingerprinting purposes have to be chosen for each
species individually. Further our PCR- RFLP results
showed the presence of Hin6I and MspI restriction
sites in two species
C.c. specularis, C. carassius
and
C.c. communis, C. carassius,
respectively.
Thus these
restriction enzymes can become identification
markers for these two species. These findings
underline the fact that mtDNA segments are more
appropriate for population studies. Our results do
showed intraspecific variations using the PCR-RFLP
of Cyt b gene. The mean nucleotide diversity of the
two species was 1.0000±0.0113 and the value of mean
haplotype diversity was 0.0000±0.0000. As expected
the value for nucleotide diversity were similar
(1.0000±0.0113) for all the species. The result also
reported by Nei (1973) evaluated inter population
polymorphism through the calculation of means of the
differences between the haplotype pairs in the sample
and of nucleotide (π) and haplotype (
h
) diversity (Nei,
1973, 1987). It is evident that the low number of
nucleotide substitutions between the haplotypes
determined the low indices of genetic diversity
(1.0000±0.0113) in the sample of cyprinid species.