INTRODUCTION
The
Tasar silkworm, A. mylitta has rich genetic resource as 44 ecoraces,
however, the Tasar culture, an important co-discipline of applied forest
biology, needs special understanding and addressing towards breeding
perspective to promote the sustainable utilization of this precious natural
resource (Reddy et al., 2009). Backcrossing
is a well known and long established breeding plan where a character is
introgressed from a domesticated or wild relative donor parent into the genomic
background of a recurrent parent, which progress better with selection of
genetically diverged parental breeds. The backcross breeding of silkworm using
parents with preferred traits and selection in subsequent generations offer
superior varieties (Reddy et al., 2009).
Assessment of genetic diversity is essential for efficient management
and conservation of any animal genetic resources in gene banks. Since, SSRs are
co - dominant markers and can reveal multiple alleles at a single locus and
been extensively used in the diversity analysis of animal and plant system. This
present work undertaken characterizes the ecoraces of Tasar silkworm,
A.
mylitta,
from different parts of tropical forest
zones, as basis for identification and genetic diversity among the tasar
populations. Based on these reports, a comprehensive breeding programme could
be evolved to conserve the dwindling population of Tasar silkworm, A.mylitta, Andhra local ecorace.
The present investigation is
an attempt to study the genetic proximity of the ecoraces of A. mylitta,
bring about an idea of breeding of Andhra local ecorace with other ecoraces,
without losing its beneficial commercial characters and suggest methods to
overcome its weaknesses and as a strategy to conserve the
dwindling population, which is on the brink of extinction. Its conservation
focused in the lines of carrying out hybridization with more than viable
variety. The selection of parental strains for a breeding program is a based on
economically desirable quantitative traits of the parental ecotypes.
MATERIAL AND METHODS
The parental stocks of ecoraces viz., Andhra Local and Daba
TV of Tasar Silkworm A. mylitta raised during the seed crop rearing
season, July-August. The Andhra local and Daba TV seed cocoons were collected
from RTRS (Warangal district) and Telangana State Silk Board (Chennur Mandal,
Adilabad district) respectively. They arranged in the form of garlands in grange
chambers. The disinfection of room is using with 2% formaldehyde, prevented
by arranging suitable nylon net, ventilated grainage chambers in the laboratory.
The date of emergence of each of the ecorace (male/female) noted. The male and
female moths emerged out of Non - diapause cocoon stocks of the above divergent
geographic ecoraces used for the study.
Isolation
of Genomic DNA
Genomic DNA was extracted by the
use of the phenol-chloroform method modified by Nagaraja & Nagaraju, (1995).
SSR
based molecular analysis of polymorphic study on the different ecoraces of Antheraea
mylitta D.
Simple sequence repeats (SSR)
otherwise known, as microsatellites are randomly repeated DNA sequence motifs
present in eukaryotic genomes. SSR markers became very popular because they are
highly reproducible, multi-allelic and Co dominant markers
The reaction mixture is
giving a momentary spin for thorough mixing of the cocktail components. Then
0.2ml PCR tubes are loaded in a thermal cycler (Research Master Cycler PTC 200,
Eppendorf) Table 3.
The thermal cycler programmed as
follows
Profile 1: 94°C for 10 min - Initial
denaturation
Profile 2: 94°C for 30 sec - Denaturation
Profile 3: 48-60°C for 35sec - Annealing
Profile 4: 72°C for 45 sec - Extension
Profile 5: 72°C for 10 min - Final
extension
Profile 6: 4°C for infinity to hold the
sample.
Profile 2, 3 and 4 programmed to run for
35 cycles.
Phylogenetic Study of different
ecoraces of Tasar Silkworm, Antheraea mylitta D.
The Phylogenetic
relationship among tasar ecoraces was analyzed by generating the Phylogenetic
tree by (Nei, 1972) genetic
distance using UPGMA analysis through POPGENE software 1.32 version (Yeh & Boyle, 1999).
The
PCR amplified bands scored visually by different ecoraces of A. mylitta because of their presence (1)
or absence (0). The scores obtained were then pooled for constructing a single
data matrix, which was used for estimating the proportion of polymorphic loci, (Yeh et al., 1999) gene diversity
(h), gene flow (Nm), coefficient of gene differentiation GST, (Nei, 1978) unbiased
genetic distance (D). Significant test and construction of a UPGMA (Un weighted
Pair Group Method of Arithmetic Means) dendrogram among populations were
carried out by using POPGENE version 1.32 (Yeh et al., 1999) computer
program. Band sharing based intra- population similarity indices (S1) were
calculated for all possible comparisons according to the following formula:
Similarity index (S1) = 2NAB / (NA + NB).
Scoring
for Co-dominant markers (SSR)
With co-dominant
markers, such as allozymes, RFLP and SSR, each recognizable allele at a given
locus is ordinarily associated with a single band at a unique position in the
gel. Thus, in the case of diploid organisms for a given locus, a homozygote
will have one band and a heterozygote will have two. Null alleles (no band)
rarely seen. In addition, if there are multiple alleles per locus, as expected
for SSRs, which are highly variable, the total number of bands expressed by all
the individuals in a sample will likely be much greater than the number of loci
involved.
In
the profile of dendrograms for SSR using popgene 1.32., the level of
polymorphism expressed as the percentage of all loci that are polymorphic. It
also gives details about no. of alleles, gene flow, genetic distance, gene
diversity, etc. Genetic Distance (D) Genetic distances designed to express
the genetic differences between two populations as a single number. If there are
no differences, the distance sent to Zero, whereas if the population has no
allele in common at any locus the distance may be set equal to its maximum
value, 1. The genetic distance (D) was calculated by POPGENE software (Yeh et al., 1999) using (Nei, 1972) standard
genetic distance equation.
RESULTS
The Instar wise average Temperature
(°C) and its standard deviation of Tasar silkworm Antheraea mylitta (F1and
F2 Hybrid) were as follows. F1 Hybrid: 32.21 ± 0.966 (S.D), 32.22 ± 2.108
(S.D), 30.14 ± 1.463 (S.D), 31.16 ± 1.834 (S.D), 29.20 ± 2.592 (S.D), and F2
Hybrid: 28.50 ± 1.322 (S.D), 28.91 ± 0.948 (S.D), 28.35 ± 0.748 (S.D), 28.78 ± 0.9063
(S.D), 29.37 ± 0.882 (S.D), for I, II, III, IV and V instars respectively. The
instar wise average Relative Humidity (%) and its standard deviation for F1
Hybrid were 86.78 ± 6.619 (S.D), 86.11 ± 6.050 (S.D), 88.42 ± 7.390 (S.D),
90.16 ± 3.544 (S.D) and 90.50 ± 4.377 (S.D) and F2. Hybrid were 93.14 ± 2.672
(S.D), 91.50 ± 4.764 (S.D), 94.00 ± 2.516 (S.D), 92.28 ± 4.644 (S.D) and 87.58 ±
3.648 (S.D) in I, II, III, IV and V instars respectively (Table 1). The no of dfls
of Tasar silkworm Antheraea mylitta (F1 and F2 Hybrid) were 02 and 01;
the mortality was 109 and 54 i.e., 33.5% and 41% respectively, and
cocoon yield was 80 and 2 Andhra local parental, Daba-TV parental, F1 Hybrid, were
found to have a genetic distance of 0.2126, 0.1629, and 0.1920 with that of F2
Hybrid. The dendrogram produced by UPGMA of Nei’s genetic distance for all
populations (4 ×13 = 52) is presented on figure 3 (Phylogenetic tree of SSR), in
(Table 4).
A
summary of genetic variation statistics for all loci depicted in the (Table 7)
indicated. The average number of alleles observed was for 1.7143 ± 0.4688,
1.6429 ± 0.4972, 1.7857 ± 0.4258, and 1.5714 ± 0.5136, for ALP, DTVP, F1
Hybrid, and F2 Hybrid respectively, the average number of effective alleles
were 1.3828 ± 0.3748, 1.3417 ± 0.3819, 1.4205 ± 0.3394, and 1.2599 ± 0.2728
respectively, while average for the total populations was 1.5555 ± 0.4047 when
all
populations were taken together, Genetic diversity (h) 0.2290 ± 0.1930, 0.2031
± 0.1979, 0.2574 ±0.1774 and 0.1724 ± 0.1701. An alternative approach for
calculating the within population variation is Shanon’s diversity index which
does not assume Hardy-Weinberg equilibrium. Average Shannon’s diversity index
was 0.3499 ± 0.2701, 0.3106 ± 0.2786, 0.3934 ± 0.2491, and 0.2707±0.2578 ALP, DTVP,
F1 Hybrid, and F2 Hybrid respectively (Table 7). The genetic diversity in the
four populations presented in (Table 7). The total genetic diversity (Ht) was
0.2290 ± 0.1930, 0.2031 ± 0.1979, 0.2574 ± 0.1774 and 0.1724 ± 0.1701.
Within-population sample genetic diversity (Hs) of ALP, DTVP, F1 and F2 Hybrids
were as 0.229 ± 0.1930, 0.2031±0.1979, and 0.2574 ± 0.1774 and 0.1724 ± 0.1701.
While overall Gene differentiation (Gst) was 0.3013 (Table
7).
Table 1. Instar-wise Temperature (°C), Relative humidity (%) of F1 and F2 hybrids of Tasar Silkworm Antheraea mylitta D.
|
Instar
|
Temperature (°C)
|
Relative Humidity (%)
|
|
F1
|
F2
|
F1
|
F2
|
|
I
|
32.21 ± 0.966
|
28.50 ± 1.322
|
86.78 ± 6.619
|
93.14 ± 2.672
|
|
II
|
32.22 ± 2.108
|
28.91 ± 0.948
|
86.11 ± 6.050
|
91.50 ± 4.764
|
|
III
|
30.14 ± 1.463
|
28.35 ± 0.748
|
88.42 ± 7.390
|
94.00 ± 2.516
|
|
IV
|
31.16 ± 1.834
|
28.78 ± 0.9063
|
90.16 ± 3.544
|
92.28 ± 4.644
|
|
V
|
29.20 ± 2.592
|
29.37 ± 0.882
|
90.50 ± 4.377
|
87.58 ± 3.648
|
*The
values expressed in terms of Standard Error of the mean.
Table 2. Rearing
performance of F1 and F2 hybrids of Tasar silkworm, Antheraea mylitta D in
2015 (for 1 DFL = 150 eggs).
|
S.No.
|
Hybrid
|
No. of dfls
|
Mortality
|
Percent Mortality
|
Cocoon yield
|
% Cocoon yield
|
|
1
|
F1
|
2
|
109
|
33.5%
|
80
|
26
|
|
2
|
F2
|
1
|
54
|
41%
|
26
|
17.3
|
Table 3. PCR amplification of DNA with SSR primers (The cocktail for PCR
amplification for respective SSR fragments is prepared. The reaction mixture (10μl)
contains).
|
PCR products
|
Amysat 005
|
Amysat 007
|
Amysat 024
|
Amysat026
|
Amysat033
|
Amysat034
|
Amysat036
|
|
Genomic DNA(10ng/µl)
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
|
10xPCR buffer
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
|
25mM MgCl2
|
0.6 µl
|
0.6 µl
|
0.6 µl
|
0.6 µl
|
0.6 µl
|
0.6 µl
|
0.6 µl
|
|
1Mm dNTPs
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
|
Forward primer
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
|
Reverse primer
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
1.0 µl
|
|
Taq DNA
polymerase
|
0.1µl
|
0.1µl
|
0.1µl
|
0.1µl
|
0.1µl
|
0.1µl
|
0.1µl
|
|
MQ
|
4.3 µl
|
4.3 µl
|
4.3 µl
|
4.3 µl
|
4.3 µl
|
4.3 µl
|
4.3 µl
|
Table 4. Nei's original measures of genetic identity and genetic
distance.
Table
5. Calculation of mean genetic distance
of parental and hybrid ecoraces of Tasar Silkworm Antheraea mylitta D (as calculated from the Table 4).
|
Sl. No.
|
Ecoraces
|
Mean values
|
|
1.
|
Andhra local parent
|
0.1473
|
|
2.
|
Daba-TV parent
|
0.1883
|
|
3.
|
F1 Hybrid
|
0.1891
|
Table 6. Polymorphism of
Hybrid populations of Antheraea mylitta D, as revealed by phylogenetic analysis
based on SSR primers.
|
Ecorace
|
Place of collection
|
Number of polymorphic loci
|
Percentage of polymorphic loci%
|
|
Andhra
local (Parental)
|
RTRS,Warangal,
Telangana
|
10
|
71.43
|
|
Daba
TV (Parental)
|
Adilabad,
Telangana
|
9
|
64.29
|
|
F1
Hybrid
|
K.U.Warangal,
Telangana
|
11
|
78.57
|
|
F2
Hybrid
|
K.U.
Warangal, Telangana
|
8
|
57.14
|
Table.7. Nei’s Analysis
of gene diversity in subdivided populations.
|
S.No.
|
Ecoraces
|
Na
|
Ne
|
h
|
I
|
Ht
|
Hs
|
|
1
|
Andhra
local Parent
|
1.7143± 0.4688
|
1.3828 ± 0.3748
|
0.2290 ± 0.1930
|
0.3499 ± 0.2701
|
0.2290±
0.1930
|
0.2290 ± 0.1930
|
|
2
|
Daba
TV Parent
|
1.6429±
0.4972
|
1.3417±
0.3819
|
0.2031 ±
0.1979
|
0.3106 ± 0.2786
|
0.2031±
0.1979
|
0.2031±
0.1979
|
|
3
|
F1
Hybrid
|
1.7857±
0.4258
|
1.4205±
0.3394
|
0.2574 ±
0.1774
|
0.3934±
0.2491
|
0.2574±
0.1774
|
0.2574± 0.1774
|
|
4
|
F2
Hybrid
|
1.5714± 0.5136
|
1.2599± 0.2728
|
0.1724±
0.1701
|
0.2707±
0.2578
|
0.1724±
0.1701
|
0.1724± 0.1701
|
Population
genetics parameters for the 11 populations of hybrid Populations of A.mylitta D. Population - wise data on the number of alleles (Na), number of
effective alleles (Ne), h = genetic diversity; I = Shannon Index, Ht = Total
genetic diversity, Hs = Sample genetic diversity, Gst = Gene differentiation,
Nm= Gene flow.
Figure
1.
Moths of Tasar Silkworm, Antheraea
mylitta D., Andhra local and
DabaTV ecoraces.
|
|
|
|
|
|
|
Andhra local parental : Lanes1–10 represent different strains of Andhra
local ecorace
Daba TV parental : Lanes
1–10 represent different strains of Daba TV ecorace
F1 Hybrid : Lanes 1–11 represent
different strains of Daba BV ecorace
F2 Hybrid : Lanes 1–10 represent different strains of Modal ecorace
M: 50
bp DNA Ladder
Note: Primer Amysat 026; Fragment
size is 142 bp
|
Figure 2. SSR profiles generated from genomic DNA of 11 strains from different
individuals of (Andhra local parental, Daba- TV parental, F1 Hybrid, and F2
Hybrid) Populations of Tasar Silkworm, A.mylitta D using the primer Amysat026.
Figure
3. UPGMA dendrogram depicting genetic
diversity of
Tasar Silkworm, A. mylitta D genotypes were obtained by PCR-SSR marker data (POPGENE
version 1.32).
DISCUSSION
In India, rearing of
silkworm hybrids is in vogue to get better cocoon yield. Since bivoltine
silkworm hybrids cannot survive the harsh climatic conditions in the tropics,
especially during the summer and rainy months, hardy tropical silkworm hybrids
need to be developed (Srivastava
et al., 2009).
The
DNA isolation and the quantification of the 1-11 samples of Tasar Silkworm, A.
mylitta D Parental
ecoraces (Andhra local, Daba TV, F1 and F2)
revealed an ideal concentration of DNA ng / µl of the genomic sample i.e., between 1.8 – 1.9 in 260 / 280
ratio) checked against 1 kb standard DNA ladder was obtained. It been
observed that DNA has been isolated without any protein or any other
contamination and was used for further studies in PCR and phylogenic analysis. Genetic
characterization of 4 populations viz.,
parental ecoraces (Andhra local, Daba TV), F1 and F2 was done. Out of the 15
primer combinations tried 6 primer combinations have shown to have desired size
fragments and three primers viz.,
Amysat024, Amysat026 and Amysat034 (of allelic size 200 - 250, 130 - 500 and
150 - 400 respectively) have shown polymorphism, while three primers viz., Amysat007, Amysat033 and Amysat036
(of allelic size 200, 200 and 250 respectively) have shown monomorphism.
The
SSR amplification of 4 tasar populations (11 individuals in each, with six
primers, which generated polymorphism) using
6 SSR primers yielded a total of 335 bands, out of which 166 bands were
polymorphic (49.55%). During scoring, all the bands present in both polymorphic
and monomorphic profiles selected. The
average no. of amplicons produced per DNA sample was 2 - 6 per primers, most
of the bands observed within the range of 150-500 bp, which is in accordance
with the allelic size of the primers taken for studies (Figure 2).
Out of a total of 335 bands produced in the
4 tasar populations, 94 bands in Amysat026, followed by 67 bands in Amysat034,
52 bands in Amysat024, 44 bands in Amysa033, 42 bands in Amysat007 and 36 bands
in Amysat036 primers were generated. Among
the 14 alleles, 11 alleles have shown polymorphism with a minimum of 2 and a
maximum 4 bands in each lane.Amysat026 primer generated a total of 94 bands,
out of which 30 bands belong to Andhra local. The no. of alleles produced 5 in Amysat026
followed by3 in Amysat024 and 1 each in Amysat007 and Amysat033 primers. From
the above observations, it can to see that Amysat026 can consider as an
effective SSR marker to identify the tasar ecotypes. From Table 4 and 5 on
genetic distance, it can be seen that F1
Hybrid
(mean value = 0.1891) shows higher genetic distance
among the other three populations (i.e.,
Andhra local parent and Daba -TV parent), which implies that F1 Hybrid is genetically
distant from other ecoraces. It observed that the lowest genetic distance found
in ecorace Andhra local parent (0.1473).
In the present investigation, screening of genomic
DNA from 11 individuals of 4 populations using 6 SSR primers yielded several
reproducible amplicons. The average no. of amplicons produced per DNA sample
were 2 - 6 per primers, with sizes ranging from 130-500 bp. The percent
polymorphism was 86.16% in Amysat 026, whi1e it was 76.11 Amysat034
and 65.38% Amysat024 SSR primers (Figure 2). In the present studies, the
germplasm collected from Andhra local,
Daba -TV and their hybrids F1 and F2 displayed variable genetic
polymorphism and was found to be highest in F1Hybrid population of Telangana
(78.57%), followed by Andhra local (71.43%) of Telangana and Daba TV (64.29%)
and F2 (57.14%) in Table 6.
The order of genetic closeness may summarize
as follows:
Daba TV parent is genetically
less close than F1 Hybrid. Andhra
local parent
is genetically less close than Daba TV parent
is.
Andhra local parent and Daba TV parent found to be close within the
populations according to phylogenetic tree. Earlier, studies using ISSR markers to analyze
the intra-race diversity in Daba (Kar et al., 2005) and Raily (Srivastava et al.,
2009) ecoraces of Antheraea mylitta
D revealed
considerable genetic differentiation across and within the populations of both
the ecoraces. In another work, using RAPD markers, genetic variation among the
different ecoraces of this moth were assessed (Saha et al.,
2008). A study of the genetic
structure of the different A. mylitta D ecoraces using polymorphic microsatellite can loci
successfully done recently Chakraborty et al.
(2015) and Renuka & Shamitha, (2016). The present work, based on
genetic analysis of 7 tasar ecoraces using SSR and ISSR generated polymorphism,
emphasized not only on the genetic closeness of Andhra local ecorace in
relation to other ecoraces, but it went further probing its compatibility to
mate with them. The resultant F1 and F2 population assessed genetically based
on the SSR primer, which reported for the first time. The molecular characterisation
and phylogenetic analysis revealed parental ecoraces (Andhra local and Daba TV)
formed one cluster, while F1 and F2 formed individual clusters. It may note
that F1 population is genetically closer to the parental cluster than that of
F2 (Figure 3).
The
present studies, which reveals a considerable improvement of commercial aspects
(Sreenivas & Shamitha, 2017) is directed towards the
conservation of Tasar silkworm, A. mylitta D, Andhra local ecorace needs further improvement of breeding
practices, to enable the viability of F1 and F2 generations. PCR - SSR based phylogenetic analysis
using popgene 1.32 in 4 tasar populations
viz., parental ecoraces
(Andhra local, Daba TV), F1 and F2), revealed that F1 Hybrid shows higher genetic distance among the other 3
populations (i.e., Andhra local parent and Daba
-TV parent),
which implies that F1 Hybrid is genetically distant from other ecoraces. It observed that
the lowest genetic distance found in ecorace Andhra local parent. Populations of Andhra local and
Daba TV (Parental ecoraces) found to be close within the populations according
to phylogenetic tree. The percent
polymorphism was 86.16% in Amysat 026, whi1e it was 76.11 Amysat034 and
65.38% Amysat024 SSR primers.
The genetic
structure of parental (Andhra local and Daba TV) and Hybrid populations (F1 and
F2) is presented in Table 7. The study clearly demonstrates that the number of
alleles (Na), number of effective alleles (Ne), genetic
diversity (h), Shannon Index (I) total genetic diversity (Ht) and
sample genetic diversity (Hs) were highest in F1 hybrid population.
This is in concurrence with the percent polymorphism, which is greater in F1
(78.57%) than that of the parental types which might be the reason for forming
an individual cluster in the phylogenetic tree. Similarly, the above population
genetics parameters were lesser in F2 hybrid populations than the parental as
well as F1 types, which resulted in the formation of individual cluster. This
is a significant observation from the breeding studies, which aimed at the
improvement of Andhra local ecorace.
According to the
recent reports, high intra population variability indicates optimum
heterozygosity (Kar et al., 2010) which is clearly demonstrated by
the F1 populations, on the other hand, the low genetic parameters of F2
populations appropriate attention towards conservation. As per the analysis of
genetic diversity amongst parental (Andhra local and Daba TV) and hybrid
populations (F1 and F2), the gene differentiation (Gst = 0.3013) and
the gene flow of (Nm = 1.1597). Clearly indicate that populations
are in the threshold of genetic differentiation, in section 4.6.1 and 4.6.2
Furthermore, the gene flow above 1.00 is the indication of mating between the
adjacent populations (Slatkin, 1987).
Nei’s analysis of gene diversity reveals the total gene diversity (Ht)
was estimated to be (0.3084 ± 0.0412), which is a higher than Hs (0.2155 ±
0.0207), while average gene diversity (Dst = Ht
- Hs) and gene differentiation were calculated to be 0.0929 ± 0.0205
and 0.3013 respectively. The values of Gst, Nm and Dst
(Dst value was not zero) suggest that some genetic differentiation
must have taken place between the populations, though at a very low pace, as
revealed by gene diversity analysis (Su et al., 2009).
The
study on the SSR based phylogenetic analysis in the parental and hybrid populations of tasar ecoraces is not so far reported. The present
investigation of hybridization based on backcross method between two
contrasting genetically variable ecoraces viz.,
of Andhra local and Daba TV was successful for two successive commercial crops.
It attributed to the fact that both these populations are inhabited in
geographically almost same altitude and comparatively lesser, when compared to
other ecoraces. It could be one of the reasons of their compatibility of mating
and production of F1 and F2 generations with quality cocoons. This is also
corroborated by the study on genetic differentiation as revealed by ISSR
markers which indicated negligible possibility of genetic mixing in high
altitude (Vijayan et al., 2006).
On one hand, Daba TV is one of the sought after ecoraces,
exploited commercially as it is more amenable to handling in the larval stages
and better suited for cocoon production with its reproductive fitness (Sinha,
2011), while on the other, Andhra local ecorace, embodied with
unique cocoon characteristics, is on the brink of extinction. It is the need of
the hour to exploit the breeding techniques and develop hybrids with these
ecoraces, which are co-existent
in this geographical zone. It is indeed much favorable if it can be standardized
in aspects like selection of parents, marker assisted breeding, optimizing
temperature and relative humidity during rearing and investigate using more
number of individuals in each ecorace, ultimately to yield cocoons without
losing their distinctive characteristics.
cONCLUSION
The present study on genetic diversity of tasar ecoraces,
selected parental ecoraces and F1 and F2 hybrid populations, as revealed by
co-dominant markers helps the prospective breeders in the development of
disease resistant, high yielding tasar populations and contribute towards their
conservation. The study needs to be explored further using more number of
primers and individuals, as genetic diversity and geographic distribution
depend on several evolutionary factors, which need to study in detail.
ACKNOWLEDGEMENTS
GS
is thankful to University Grants Commission (New Delhi), for providing funds
required to carry out the present work under Major Research Project UGC/MRP: F No. 42-528/2013 (SR), 22nd,
March 2013).
REFERENCES
Chakraborty, S., Muthulakshmi, M., Vardhini, D., Jayaprakash,
P., Nagaraju, J., & Arunkumar, K. (2015). Genetic analysis of Indian tasar
silkmoth (Antheraea mylitta)
populations. Scientific Reports, 5,
15728.
Kar, P., Srivastava, A., Sinha, M., Sinha, A., & Prasad,
B. (2010). Conservation genetics, combination of molecular biology and ecology:
tropical tasar silkworm as an example. Bioscan,
3, 627-634.
Kar, P., Vijayan, K., Mohandas, T., Nair, C., Saratchandra,
B., & Thangavelu, K. (2005). Genetic variability and genetic structure of
wild and semi-domestic populations of tasar silkworm (Antheraea mylitta) ecorace Daba as revealed through ISSR markers. Genetica, 125(2-3), 173-183.
Nagaraja, G. M., & Nagaraju, J. (1995). Genome
fingerprinting of the silkworm, Bombyx
mori, using random arbitrary primers. Electrophoresis,
16(1), 1633-1638.
Nei, M. (1972). Genetic distance between populations. The American Naturalist, 106(949),
283-292.
Nei, M. (1978). Estimation of average heterozygosity and
genetic distance from a small number of individuals. Genetics, 89(3), 583-590.
Reddy, R. M., Suryanarayana, N., Sinha, M. K., Gahlot, N. S.,
Hansda, G., Ojha, N. G., & Prakash, NBV. (2009). Silk filament progression
with backcross breeding generations in tropical tasar silkworm, Antheraea mylitta D. International Journal of Industrial
Entomology, 19(1), 187-192.
Renuka, G., & Shamitha, G. (2016). Genetic variation in
ecoraces of tropical tasar silkworm, Antheraea
mylitta using SSR markers. Journal of
Genetics, 95(4), 777-785.
Saha, M., Mahendran, B., & Kundu, S. (2008). Development
of random amplified polymorphic DNA markers for tropical tasar silkworm Antheraea mylitta. Journal of Economic Entomology, 101(4), 1176-1182.
Sinha, A.K. (2011). Variability in the Ecoraces Of Tropical
Tasar Sillkworm Antheraea Mylitta
Drury. Nature Precedings : doi:10.1038/npre.2011.6161.1.
Slatkin, M. (1987). Gene flow and the geographic structure of
natural populations. Science, 236(4803),
787-792.
Sreenivas, M., & Shamitha, G. (2017). Comparative study on
rearing performance, larval and post-cocoon characters of Tasar silkworm, Antheraea mylitta Drury ecoraces (Sukinda, Daba TV and Andhra local).
Journal of
Entomology and Zoology Studies. 5(2): 1348-1356.
Srivastava, A.
K., Kar, P. K., Sinha, R., Sinha, M. K., & Vijayaprakash, N. B. (2009).
Assessment of genetic diversity in different populations of raily ecorace of
indian tasar silkworm, Antheraea mylitta Drury
using ISSR markers. International Journal
of Industrial Entomology, 19(2), 249-253.
Su, W., Michaud,
J., Runzhi, Z., Fan, Z., & ShuAng, L. (2009). Seasonal cycles of
assortative mating and reproductive behaviour in polymorphic populations of Harmonia axyridis in China. Ecological Entomology, 34(4), 483- 494.
Vijayan, K.,
Anuradha, H., Nair, C., Pradeep, A., Awasthi, A., Saratchandra, B., Urs, S.R.
(2006). Genetic diversity and differentiation among populations of the Indian
eri silkworm, Samia cynthia ricini,
revealed by ISSR markers. Journal of
Insect Science, 6(1), 1-11.
Yeh, F., Yang,
R., & Boyle, T. (1999). Popgene version 1.32: Microsoft Windows based
freeware for population genetic analysis. Quick User Guide University of Alberta Press, 1-28.
