Genetic characterization of Metapenaeus affinis (H. M. Edwards, 1837) using RAPD markers

Genetic structure of four populations of Metapenaeus affinis from Maharashtra, Orissa, Kerala and Tamil Nadu in India was studied using RAPD markers. Five selective primers provided distinct and consistent RAPD profiles in all the four populations. The bands in the range 225–1,900 bp were scored for consistent results. The RAPD profiles generated by all the five primers revealed varying degrees of polymorphism, ranging from 25.00% (primer E-03) to 65.00% (primer E-06). Nei’s (Nei M, Natl Acad Sci Proc USA 70:3321–3323, 1973) genetic diversity (h) among the four populations varied from 0.2565 ± 0.2146 (Orissa population) to 0.3576 ± 0.1897 (Maharashtra population).     

Length–weight relationships of Barilius bendelisis (Hamilton, 1807), Barilius shacra (Hamilton, 1822) and Barilius barna (Hamilton, 1822) from Manas River in Assam,India

The present study describes the length–weight relationships (LWRs) for three ornamental hill stream fish species from the Manas River in Assam, India, namely, Barilius bendelisis (Hamilton, 1807), Barilius shacra (Hamilton, 1822), and Barilius barna (Hamilton, 1822). Fishes were collected on a monthly basis from March 2015 to February 2016 with cast nets (270 cm, 1.2 cm) and gillnets (7,500 × 130 cm, 5 cm). This is the first information on LWR data for two of the species.     

Genetic relationships among NE Turkish Lilium L. (Liliaceae) species based on a random amplified polymorphic DNA analysis

Random amplified polymorphic DNA (RAPD) fingerprinting was used to study species boundaries in six closely related NE Turkish Lilium (Liliaceae) taxa of the section Liriotypus. The investigated taxa were L. ciliatum, L. akkusianum, L. ponticum, L. kesselringianum, L. armenum, and L. szovitsianum. Of the 108 primers screened, 11 provided polymorphic and reproducible bands. A total of 93 polymorphic bands were scored for 122 individuals from 18 populations of the six Lilium taxa and principle coordinate analysis and neighbour-joining cluster analysis based on these RAPD profiles were performed. The results demonstrate a clear distinction between the two species L. ciliatum and L. akkusianum, and the other four species. While populations of the two species groups are found to be allopatrically distributed, the two species groups overlap in their geographical ranges. Analysis of molecular variance (AMOVA) indicated that nearly half of the total molecular variance is found within the individual populations and that the molecular variance among species is as high as the variance within the individual species, indicating that genetic differentiation of the species is rather weak.     

Genetic Characterization of <Emphasis Type="Italic">Barilius barna</Emphasis> (Hamilton, 1822) in the Teesta River of Sub-Himalayan West Bengal,India, Through RAPD and ISSR Fingerprinting

Genetic characterization of Barilius barna, an economically important freshwater fish in the Indian scenario, is unexplored in the sub-Himalayan Dooars region of West Bengal, India. This study is the first attempt to characterize the genetic architecture of Barilius barna from the Teesta river of this region. We have studied loci polymorphism, genetic diversity, Shannon’s information index and the measure of evenness in the two populations of this river through ten RAPD and seven ISSR primer-based PCR amplifications. The result showed 89.52 and 82.21% polymorphisms in RAPD and ISSR amplification respectively. The Nei’s genetic diversity and Shannon’s information index varied from 0.172 ± 0.189SD to 0.293 ± 0.164SD and 0.265 ± 0.268SD to 0.445 ± 0.220SD respectively, which indicated low level of genetic variation. AMOVA revealed significant level of variance within the population and gene flow between the populations. Low levels of genetic variation and moderate to high levels of genetic relatedness were found in the studied populations. Expectedly, the populations were genetically not very distant from each other, as evident from the Nei’s unbiased measure of genetic distance and identity. As the species is commercially important and the region is located in the sub-Himalayan region, the management and proper rehabilitation of this ichthyofauna in the wild is urgently required. Our results may serve as a guideline for adopting such management decisions.     

Phenotypic divergence and molecular characterization of two sympatric species of genus Barilius (Hamilton) from the River Chenab of Western Himalaya

Barilius bendelisis and Barilius vagra inhabit mostly in the spring‐fed streams of Indian Himalaya and constitute an important portion of protein diet of the rural human population residing near the stream banks. In spite of large aquaculture importance of this genus very less morphometric and molecular studies have been conducted. This study addresses morphometric and molecular characterization of B. bendelisis and B. vagra by using truss analysis and mitochondrial COI gene. A total of 293 samples of Barilius bendelisis and 127 samples of Barilius vagra were collected from three different sites of Chenab river basin from March 2015 to April 2017. 14 landmarks were used to measure 90 truss measurements from digital images of specimens using network of three softwares, tpsUtil, tpsDig2 and PAST. A total of 89 distance variables exhibited significant differences among the populations. The principal component analysis generated ten components explaining 89.164% of total variance among six populations of B. bendelisis and B. vagra. 89.0% of individuals of B. bendelisis and B. vagra were classified into their original groups. The Phylogenetic analysis with other published COI sequences revealed distinct nature of these two species. The study may aid in taxonomic identification and phenotypic divergence which will be helpful in proper conservation and management of these species of hill trout in India.     

Genetic diversity analysis of maintainer and restorer accessions in upland cotton (Gossypium hirsutum L.)

Genetic diversity amongst 91 upland cotton accessions (50 maintainer, ‘B’ and 41 restorer ‘R’ lines) and three wild species viz., G. aridum, G. thurberi and G. anomalum was analyzed using SSR and RAPD markers. A total of 53 primers (30 SSR and 23 RAPD) were sampled for screening 94 accessions, of which 26 SSR and 17 RAPD primers were polymorphic. Average polymorphism detected by SSR, RAPD and SSR + RAPD markers was 72.5, 62 and 66.66 per cent, respectively. A unique marker CIR-200₂₆₀ that distinguishes G. thurberi from all upland accessions has been identified. Similarity coefficient values within and between B and R lines ranged from 0.65–0.95, 0.61–0.98 and 0.53–0.93 for SSR and 0.72–0.98, 0.73–0.97 and 0.69–0.98 for RAPD markers. UPGMA cluster analysis was consistent with the pedigree and genotypic background of the accessions. RAPD and SSR matrices showed significant positive product moment correlation (r?=?0.93 and 0.92) with the RAPD + SSR combined data matrix, respectively. The result indicates a moderate level of genetic diversity in B and R accessions of upland cotton. Genetically diverse combinations were identified to further evaluate heterotic performance. The maintainer, AKH-108, AKH-118 and AKH-2173; and restorer AKH-31 and AKH 4943 accession were identified as most distinct and divergent, could be used as candidate parental genotypes in hybrid and varietal development programme and also development of mapping population for trait mapping in cotton.     

Biochemical and Molecular Characterization of Some Centaurea Species Growing in the Eastern Anatolia Region of Turkey

Random amplified polymorphic DNA (RAPD) and fatty acid (FAME) profiles were used to examine phenotypic and genetic relationships among 16 Centaurea species growing wild in the eastern Anatolia region of Turkey. Thirteen decamer primers were used to examine polymorphism. According to the RAPD results, 99 amplicons in the size range of 50–1000 bp were produced from 13 primers in 16 Centaurea species. Genetically four distinct groups were determined among the species of Centaurea, which represents high genetic variation. In the 16 species, 14 fatty acids were determined according to FAME results. Both FAME and RAPD results showed that C. virgata is genetically different from other species. The differences in the composition of fatty acids among Centaurea species suggest that fatty acid profiles could be used to differentiate among some of these species. Results of this study show that RAPD and FAME analyses are consistent.     

Genotyping of Ganoderma species by improved random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) analysis

Species of Ganoderma are used in traditional medicines. An improved random amplified polymorphic DNA (RAPD) analysis, where the RAMP time is prolonged, has been used to characterize the genetic variation in some well known species of Ganoderma. The DNA materials were collected from ten Ganoderma strains, amplified with randomly selected 24 RAPD primers and evaluated by agarose gel electrophoresis. A cluster dendrogram was constructed for genetic analysis on the basis of amplification results. The improved RAPD amplified DNA with consistent and clear banding patterns. A total of 316 bands were found with 93% polymorphism. There was a significant genetic distance between the different strains of Ganoderma, with an index of similarity coefficient in the range of 0.52–0.74. The inter-simple sequence repeat (ISSR) analysis of the Ganoderma DNA samples showed similar trend results to the RAPD analysis with 0.49–0.81 similarity coefficients. This study reports the high level of genetic differences between different species or strains of a single species of Ganoderma and confirms the significance of the improved RAPD method in genetic characterization of organisms. Therefore, the improved RAPD combined with ISSR techniques might be used for the genetic characterization of organisms.     

Inter and intraspecific genetic diversity (RAPD) among three most frequent species of macrofungi (Ganoderma lucidum,Leucoagricus sp. and Lentinus sp.) of Tropical forest of Central India

In present study seven RAPD primers were used to access the diversity within and among twelve populations of three mushroom species Ganoderma lucidum, leucoagaricus sp. and Lentinus sp. Total of 111 bands were scored by 7 RAPD primers in 30 accessions of three mushroom species collected from different sampling sites of central India. Total 111 bands were generated using seven primers which were F-1, OPG-06, OPC-07, OPD-08, OPA-02, OPD-02, OPB-10. All 111 bands were polymorphic in nature (100%). Therefore, it revealed that the used primers had sufficient potency for population studies and 30 accessions had higher genetic differences among each other. In best of the knowledge, this is the first report, which accesses the genetic diversity between three mushroom species (Gd Ganoderma lucidum, Lg Leucoagaricus sp., Ls Lentinus). The polymorphic percentage ranged from 3.60 to 23% within twelve populations, while polymorphic percentage among group was 40.56, among population within groups was 41.12 and within population was 18.32. This indicated that the genetic diversity within the population was very low, but slightly higher in the populations of three species. Among three groups representing Gd., Lg and Ls, Among populations within groups shown highest percentage of variation (Pv?=?41.12) while within populations, the lowest percentage of variation (18.32) was observed. This result also support that the highest genetic variation was present among groups in comparison to among the population within a species and lowest genetic variation was observed within the population.     

Confirmation of relationships among Boesenbergia (Zingiberaceae) and related genera by RAPD

The relationships among 19 accessions of Zingiberaceae belonging to 11 species of Boesenbergia, six species of Kaempferia, and two species of Scaphochlamys from Southern Thailand were studied using random amplified polymorphic DNA (RAPD) profiles from leaf tissue samples. The RAPD was carried out using 10 random decamer arbitrary primers. Amplification occurred in five out of 10 tested primers (OPAM-01, OPAM-03, OPAM-12, OPB-14, OPZ-03). Total of 53 amplified bands were observed. Data obtained from the RAPD fingerprints from the samples clarified some doubts in morphological classification. The data were analyzed for the Nei and Li's Dice similarity coefficient for pair-wise comparison between individual samples and the distance matrix. The dendrogram resulting from cluster analysis, UPGMA and a principal component analysis of the RAPD result confirms a higher degree of relationship between Boesenbergia and Scaphochlamys than between Boesenbergia and Kaempferia.     

Lack of genetic variation of an invasive clonal plant Eichhornia crassipes in China revealed by RAPD and ISSR markers

Random amplified polymorphic DNAs (RAPDs) and inter-simple sequence repeats (ISSRs) markers were used to analyze genetic structure of six populations of invasive plant Eichhornia crassipes that were sampled from its introduced regions in Southern China. Using 25 RAPD primers and 18 ISSR primers, 172 RAPD bands and 145 ISSR bands were produced respectively. But no polymorphic band was detected either within population or among populations by both markers, indicating the genetic diversity of E. crassipes in Southern China is extremely low, and all populations most likely consist of the same genotype. This study suggested that some other adaptability related factors, other than the genetic diversity, are responsible to the E. crassipes rapid expansion in China.     

Life history variation between species of the relictual genus Borderea (Dioscoreaceae): phylogeography, genetic diversity, and population genetic structure assessed by RAPD markers

The genus Borderea consists of two species, B. pyrenaica and B. chouardii, taxa which have been previously considered as conspecific due to their overall close morphology. These two sole species of the rare genus of Dioscoreaceae are endemic to the Pyrenees (Spain, France). This mountain range likely operated as a refugium for these plants during the last glaciations. B. chouardii is only known from a single population in the Spanish Prepyrenees and has been classified as at risk of extinction in the Red List of Endangered Species (IUCN); B. pyrenaica shows a narrow distribution range in the central Pyrenees and Prepyrenees. We analysed genetic variation, population structure and differentiation in these two taxa using RAPD markers. Our study was conducted on the same seven populations for which very low levels of genetic differentiation were detected previously through allozyme analysis. By contrast, high levels of genetic variability were detected through the RAPD hypervariable markers. Twelve RAPD primers produced 112 distinct bands in the 397 surveyed individuals, totalling 395 different RAPD phenotypes. Only four bands were monomorphic across all samples of Borderea, whereas 21 of the polymorphic bands were species‐specific (20 for B. chouardii, and one for B. pyrenaica). The largest genetic distances were those between the B. chouardii and the B. pyrenaica phenotypes. An analysis of molecular variance showed greater variance between groups (B. chouardii vs. B. pyrenaica, 76.08%) than within groups (3.60%). RAPD band specificity, phenotypic distances, and the partitioning of variance all support the taxonomic separation of the two species. Statistical evaluation of within‐ and among‐population RAPD genetic variability in B. pyrenaica showed that genetic variability was higher within populations (>80%) than among them. No clear pattern of RAPD differentiation could be observed among the six studied populations of this taxon though slight differences in genetic diversity could be observed in the more isolated Prepyrenean populations compared with the more widespread Pyrenean ones. These results suggest a recent postglacial origin of the present B. pyrenaica populations. © 2003 The Linnean Society of London, Biological Journal of the Linnean Society, 2003, 80 , 483–498.     

Assessment of random amplified polymorphic DNA (RAPD) for determining the origin ofYoungia koidzumiana Kitamura (compositae)

We determined the parental species ofYoungia koidzumiana (a natural interspecific hybrid) using PCR and arbitrary 10-mer primers to generate random amplified polymorphic DNA (RAPD) markers. These markers, generated by three primers, were sufficient to distinguishYoungia sonchifolia, Youngia denticulata, Youngia chelidoniifolia, andY. koidzumiana. The electrophoresis profiles of the amplified products from each of the four species were then compared. Three primers produced a total of 42 scorable markers; nine were specific markers forY. denticulata andY. chelidoni-ifolia. The length of the amplified DNA fragments ranged from 370 to 2500 b p. The three primers revealed polymorphic bands, which were indicators of the parental species ofY. koidzumiana. These bands showed a combination of specific profiles forY. denticulata andY. chelidoniifolia. Our results also were comparable to the data obtained for flowering times, floret numbers, and chromosome numbers of the four species. Therefore, we suggest thatY. koidzumiana is a hybrid betweenY. denticulata andY. chelidoniifolia}, and that RAPD markers are well suited for assessing the origins of plant species.     

Comparative Evaluation of RAPD and AFLP Based Genetic Diversity in Brinjal (Solanum melongena)

The present investigation was carried out with an objective of evaluating genetic diversity in brinjal (Solanum melongena) using DNA markers. A total of 38 brinjal accessions including one wild-species, Solanum sisymbrifolium were characterized using random amplified polymorphic DNA (RAP D) and amplified fragment length polymorphism (AFLP) techniques. Out of 45 primers employed to generate RAPD profiles, reproducible patterns were obtained with 32 primers and 30 (93.7%) of these detected polymorphism. A total of 149 bands were obtained, out of which 108 (72.4%) were polymorphic. AFLP analysis was carried out using four primer combinations. Each of these primers was highly polymorphic. Out of 253 fragments amplified from these four primer combinations, 237 (93.6%) were polymorphic. The extent of pair-wise similarity ranged from 0.264 to 0.946 with a mean of 0.787 in RAPD, in contrast to a range of 0.103 to 0.847 with a mean of 0.434 in AFLP. The wild species clustered separately from the brinjal genotypes. In the dendrogram constructed separately using RAPD and AFLP markers, the brinjal genotypes were grouped into clusters and sub-clusters, and the varieties released by IARI remained together on both the dendrograms. All the 30 RAPD primers in combination and each of the four primer pairs in AFLP could distinguish the brinjal accessions from each other. AFLP was thus found to be more efficient than RAPD in estimation of genetic diversity and differentiation of varieties in brinjal.     

A contribution to characterisation of genetic variation in some natural Polish populations of Elymus repens (L.) Gould and Elymus hispidus (Opiz) Melderis (Poaceae) as revealed by RAPD markers*

To determine the relative importance of clonal growth and sexual reproduction, the Randomly Amplified Polymorphic DNA (RAPD) method was used to study genetic diversity and clonal structure of six populations of Elymus repens and four populations of Elymus hispidus from Poland. These outbreeding species are virtually self‐sterile and form widely spreading and long‐lived rhizomes. Using 12 primers, a total of 150 unambiguous RAPD fragments were amplified and scored. Results of AMOVA showed no significant genetic distinction between morphologically distinguished varieties of E. repens and E. hispidus. E. repens had slightly higher intra‐specific genetic polymorphism than E. hispidus; the percentage of polymorphic bands per population ranged from 38 to 49 and from 19 to 38 respectively. Clonal diversity measured using the Simpson diversity index (D) indicated different contributions of clonal reproduction in particular populations of E. repens (D: 0.20–0.72). Populations of E. hispidus were dominated by one or a few clones, which were generally restricted to a single population (D: 0.00–0.22). RAPD revealed that most genetic diversity resided within populations of the two studied species, suggesting that, despite their clonal character, propagation by seeds contributes considerably to reproduction of E. repens and E. hispidus.     

RANDOM AMPLIFIED POLYMORPHIC DNA (RAPD) MARKERS REVEAL CROSS-FERTILISATION IN BIOMPHALARIA GLABRATA (PULMONATA: BASOMMATOPHORA)

The freshwater, schistosome-transmitting snail, Biomphalariaglabrata is a simultaneous hermaphrodite which can reproduceby both cross-and selffertilisation, but despite the medicalimportance of this species, little is known about the matingstrategies adopted by wild-type individuals from natural populations.The identification of cross-fertilised progeny is a prerequisitefor both evolutionary studies of the relative fitness of differentreproductive strategies, and for the production of exclusivelyoutcrossed F₁ populations for genetic mapping of biologicallyimportant phenotypes, most notably, resistance to schistosomeinfection. In this study, parents, offspring and ‘syntheticoffspring’ (controls) from four families of B. glabratarecently derived from wild populations were analysed using theRandom Amplified Polymorphic DNA (RAPD) assay. RAPDs were usedbecause they reveal more genetic variation, require less tissue(juvenile snails were only 4–5 mm diameter) than proteinelectrophoresis, are less laborious than other molecular analyses,and do not require sequence data. Seven of the 19 arbitrarysequence, oligonucleotide primers used gave bands that werepolymorophic between pairs of parent snails. Scoring the offspringfor the presence/absence of these polymorphic bands showed thatall the offspring tested were the products of cross-fertilisation.This study provides the first demonstration of the applicabilityof the RAPD technique to an analysis of fertilisation in wild-typeB. glabrata. (Received 20 December 1994; accepted 27 March 1995)     

Genetic diversity within and among populations of the endangered species Taxus fuana (Taxaceae) from Pakistan and implications for its conservation

The West Himalayan yew, Taxus fuana Nan Li & R.R. Mill (Taxaceae), is an endangered species endemic to the Western Himalayas. An investigation of the genetic diversity of wild populations of T. fuana in Pakistan was undertaken. The genetic diversity and genetic structure was quantified using random amplified polymorphic DNA (RAPD) variation in 219 individuals of the 10 populations. Of the 32 universal primers screened 16 produced highly reproducible, clear RAPD bands. Using these primers, 193 discernible DNA fragments were generated, of which 164 (84.97%) were polymorphic. The statistical results indicated that there was a relatively low genetic diversity within populations (with percentages of polymorphic bands, PPB, ranging from 29.53 to 50.26%, with an average of 38.34% and a Nei's genetic diversity index (H_E) of 0.1165), and a high genetic differentiation among populations (G_ST = 0.5842, Φ_ST = 0.5685) within these populations. The gene flow (N_m) was low with only 0.3558.     

Genetic Diversity Within and Among Lepidium draba Populations from Eastern Anatolia Based on RAPD Analysis

Genetic variation and structure of six natural populations of Lepidium draba L. from Eastern Anatolia were assessed using random amplified polymorphic DNA (RAPD) markers. For RAPD analysis, 12 primers generated 218 reproducible bands across the six populations analyzed, of which 73 bands (33.3%) were polymorphic. The mean Nei’s gene diversity value for all six populations was 0.1771. Shannon’s information index varied with population (0.2278–0.3082), averaging 0.2608. Analysis of molecular variance (AMOVA) showed that genetic diversity was greater within populations (58.66%) than among populations (30.68%). In addition, the variation between groups was 10.33%. The genetic differentiation among populations (G _ST) was 0.3210, indicating that most genetic diversity occurs within populations. Gene flow (Nm) was low, at only 0.5288.     

Molecular genetic diversity of Satureja bachtiarica

Fifty-seven genotypes from eight population of Satureja bachtiarica was evaluated using fifteen ISSR and eleven RAPD markers. DNA profiling using RAPD primers amplified 84 loci, among which 81 were polymorphic with an average of 7.36 polymorphic fragments per locus. Also, using RAPD markers maximum and minimum polymorphic bands observed for Semyrom (77.38 %) and Farsan (40.48 %) populations, respectively. Semyrom population recorded the highest unbiased expected heterozygosity (0.259) and Shannon’s Indices (0.38). While, the lowest values of unbiased expected heterozygosity (0.172) and Shannon’s Index (0.245) were recorded for Eghlid and Farsan populations, respectively. On the other hand, ISSR primers produced 136 bands, from which 134 were polymorphic with an average of 9.06 polymorphic fragments per primer (98.52 %). The ISSR markers evaluation revealed that maximum and minimum polymorphic bands observed for Semyrom (66.18 %) and Farsan (31.62 %), respectively. Shahrekorud population recorded the highest unbiased expected heterozygosity (0.211) and Shannon’s Indices (0.301). While, the lowest value of unbiased expected heterozygosity (0.175) observed for Farsan and Yazd populations and the lowest Shannon’s Index (0.191) recorded by Farsan population. The overall results of the study revealed that both ISSR and RAPD markers were effective for evaluation of genetic variation of S. bachtiarica.