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Abiya Chelliah D1, Inter Simple Sequence Repeats (ISSR) markers were utilized to identify the
Selvin Samuel A1 and levels of heritable varieties and patterns of the populace structure among the five
John De Britto A2 populaces of Pteris biaurita, a natural fern in India. A comprehensive examination
was directed in three replicates at 2013-14 seasons in the Western Ghats, South
India. Five wild P. biaurita, accessions (maiden hair) were assessed for genotyping
studies. Results demonstrated a pivotal discrepancy among genotypes for they were
Institution:
characterized in view of this uniqueness in four groups by the genetic cluster
1. Department of Botany,
St. Johns College, examination. In this trial, ISSR primers amplified 63 polymorphic groups. In view of
Palayamkottai - 627 002, the genetic identity data, genotypes were figured and differed from 0.5714 to
Tamil Nadu. 0.6984. The percentage of polymorphism indicated predominant genotype that may
be utilized for the conservation of species. ISSR appeared to be an obliging marker
for prediction of genotype inside a closed group of inter specific populace in the
2. Department of Botany, investigation territory.
St. Xaviers College,
Palayamkottai - 627 002,
Tamil Nadu Keywords:
ISSR analysis, Pteris biaurita L, genetic variation, Western Ghats
Corresponding author:
Abiya Chelliah D
Email Id:
Article Citation:
Heritable variations of Pteris biaurita L. discovered by ISSR markers in the Western
Ghats of Tamil Nadu, India
Abiya Chelliah D, Selvin Samuel A and John De Britto A
Journal of Research in Biology (2016) 6(6): 2078-2084
Dates:
Received: 15 May 2016 Accepted: 31 Jul 2016 Published: 18 Aug 2016
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Table 1. Place of collection of the plants and their Table 2. ISSR primers and their sequences
accession ID S. No. Name of the primer Primer Sequences
S. No. Species Accession ID Location 1 B07 (CG)5AG
POP 1 Kothayar 2 B09 (ATG)3CA
POP 2 Gundar 3 L03 (GC)4 AT
Pteris POP 3 Thirugurankudi
1. 4 A12 (GA)6CC
biaurita
POP 4 Kodaikanal 5 A13 (GT)6CC
POP 5 Kadana dam DNA polymerase, 10 mM dNTP, 10 M primer in 1
MATERIALS AND METHODS reaction buffer that contained 10 mM Tris-HCl (pH 8.3),
Study area 50 mM KCl, 2.5 mM MgCl2, and 0.01% gelatine
Five Western Ghats regions viz., Kothayar, (Williams et al., 1990). Amplifications were performed
Gundar, Thirugarankudi, Kodaikanal and Kadana Dam in an Eppendorf Master Cycler gradient. Amplification
are chosen for the study due to the ease of use of this conditions were one cycle at 94C for 4 min, and 94C
three species unvaryingly (Table 1). Ten ISSR markers for 30 s, 55C for 45 s, followed by stepwise reduction
were veteran which showed steady amplification in five of 1C for the first five cycles, and 72C for 2 min. In
markers at five sampling sites. This has provided a clear subsequent 35 cycles, annealing temperature was
idea about the genetic diversity of Pteris biaurita in its maintained at 50C, followed by one cycle of 7 min at
species level. 72C. Amplified yields were loaded on 2% agarose gel
DNA isolation and separated in 1 TBE buffer at 75 V. The gels were
Pteris biaurita, individuals were collected from visualized under UV after staining with ethidium
the Western Ghats of Tamil Nadu, India. DNA was bromide and documented using a gel documentation and
extracted from youthful leaves using the method image analysis system. The primers used for the ISSR
described by Dellaporta et al. (1983). The DNA isolated analysis are listed out in Table 2.
was purified using Phenol- Chloroform method and the Data Analysis
purity of the DNA samples were determined using UV- The gels from ISSR investigation were
Spectrophotometer at the optical density of 260 nm and visualized at gel documentation system (Alpha Imager
1
280 nm; the DNA samples were diluted to 25 ng l for 1200). Based on the primary data (presence or absence of
PCR amplification. bands), pair wise genetic distance between the samples
ISSR amplification was calculated using NTSYS and POPGENE packages.
ISSR amplification reactions were carried out in
25-l volume containing 50 ng template DNA, 0.5 U Taq RESULTS AND DISCUSSION
Figure 1. UPGMA dendrogram of Pteris biaurita based on Neis genetic distance derived using ISSR markers
L03-7 B09-13
L03-6 B07-1
L03-5 B07-2
L03-4 B07-3
Gene Diversity
L03-3 B07-4 Mean Natural Selection Coefficient
L03-2 B07-5
L03-1 B07-6
A21-15 B07-7
A21-14 B07-8
A21-13 B07-9
A21-12 B07-10
A21-11 B07-11
A21-10 B07-12
A21-9 B07-13
A21-8 B07-14
A21-7 B07-15
A21-6 B07-16
A21-5A21-4 B07-17
A21-3 A21-2A21-1
Figure 2. Summary of genetic variation statistics for all loci in Pteris biaurita
nearness and relativity. Out of ten primers five populaces of diploid homosporous ferns are generally
uncovered steady banding pattern and in this way ruled by sexual random mating or outcrossing. Besides,
uncovered variability inside the species. gene flow from the adjoining populaces would moderate
ISSR markers were utilized to analyze genetic the broadening yet just by means of sexual proliferation.
demarcation inside and between the chose species. Associated designs have been recognized in various
Investigation of genetic variability showed all the five other ferns (Ranker, 1992; Ranker and Geiger, 2008;
populaces analyzed, sexual recombination had been the Rumsey et al., 1999; Chen et al., 2010).
greatest establishment of hereditary variety than asexual The purpose behind this hereditary variety is
proliferation (Mes, 1998; Hulst 2000; Kjolner et al., probably emerged from contrasts in the DNA content of
2006). This decision is in congruity with different past the progenitor species. This proposal is reliable with the
examinations, which have almost utilized allozyme event of diploid apomicts in different plants, including
markers to derive the mating frameworks working in taxa of the Pteris biaurita (Pravin, 2005). The clades are
normal populaces of the related and locally accessible developed through Popgene 2.1 and related ancestries
ferns. indicate brawny patterns of reticulate fruition. Higher the
Allozyme analysis by Soltis and Soltis (1987) genetic diversity of these species, more prominent is
and Ranker and Geiger (2008) demonstrated that their practicality in the earth.
Researches have brought up that this induction Chen YY, Han QX, Cheng Y, Li ZZ and Li W.
ought to be confined to close relatives, and the (2010). Genetic variation and clonal diversity of the
distinction amongst diploids and their autoploid endangered aquatic fern Ceratopteris pteridoides as
offspring (Moran, 1982; Barrington et al., 1986). Our revealed by AFLP analysis. Biochemical Systematics and
discoveries are predictable with proposals that the high Ecology, 38(6):1129-1136.
chromosome numbers, and conserved chromosome sizes
David Barrington S, Cathy Paris A and Thomas
revealed for some homosporous ferns. This has been
Ranker A. (1986). Systematic inferences from spore and
recommended to be because of a higher degree of
stomata size in the ferns. American Fern Journal, 76
consistency of chromosomes and the possible
(3):149-159.
concealment of transposable components (TEs) in
homosporous ferns (Barker, 2013; Bainard et al., 2011). Dellaporta SL, Wood J and Hicks JB. (1983). A plant
It likewise affirms with the theory that genomes DNA minipreparation: version II. Plant Molecular
size variation in homosporous ferns are driven by poly- Biology Reporter, 1(4): 19-21.
ploidisation. Our examination proves that the genome
Kjolner S, Sstad SM and Brochmann C. (2006).
evolution is happening in these plants; without a doubt,
Clonality and recombination in the arctic plant Saxifraga
given the degree of hybridization and are most likely
cernua. Botanical Journal of the Linnean Society, 152:
more broad across ferns, however, may have been to a
209217.
great extent disregarded because of the reticulate
advancement detailed in homosporous ferns; in general, Manickam VS. (1995). Rare and endangered ferns of
it appears to be likely that adjustments in genome size Western Ghats of South India. Fern Gazette; Journal of
low level of sampling. the British Pteridological Society, 15(1): 1-10.
Van der Hulst RGM, Mes THM, Den Nijs JCM and
Bachmann K. (2000). Amplified fragment length
polymorphism (AFLP) markers reveal that population
structure of triploid dandelions (Taraxacum officinale)
exhibits both clonality and recombination. Molecular
Ecology, 9(1): 18.
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