Comparative Analysis of RAPD and ISSR Markers for Characterization of Sesame (Sesamum indicum L) Genotypes

The determination of genetic differences among crop genotypes has become the primary need to grant patent and the protection of Plant Breeder Rights (PBR). In the present study RAPD and ISSR markers were employed for the characterization of 16 sesame genotypes. Twenty six RAPD and 17 ISSR primers that generated clear and reproducible banding patterns amplified 194 and 163 bands, respectively among 16 sesame genotypes. Both RAPD and ISSR primers showed maximum discrimination power, and produced putative variety specific bands, which could be used for the identification of all the sesame genotypes, individually. However, only AG and CA based ISSR primers were found effective in the discrimination of genotypes. A poor correlation was observed between the matrices produced by RAPD and ISSR primers, which might be due to the array of different sites of the genome. Though, there was greater similarity among sesame genotypes (0.78 for RAPD and 0.71 for ISSR), the observed genetic diversity (0.22 for RAPD and 0.29 for ISSR), was found effective for the characterization of sesame genotypes. It is suggested that putative variety specific RAPD and ISSR markers could be converted to Codominant sequence characterized amplified region/sequence tagged site (SCAR /STS) markers to develop robust variety specific markers.     

Comparison of RAPD,ISSR, and AFLP Molecular Markers to Reveal and Classify Orchardgrass (Dactylis glomerata L.) Germplasm Variations

Three different DNA-based techniques, Random Amplified Polymorphic DNA (RAPD), Inter Simple Sequence Repeat (ISSR) and Amplified Fragment Length Polymorphism (AFLP) markers, were used for fingerprinting Dactylis glomerata genotypes and for detecting genetic variation between the three different subspecies. In this study, RAPD assays produced 97 bands, of which 40 were polymorphic (41.2%). The ISSR primers amplified 91 bands, and 54 showed polymorphism (59.3%). Finally, the AFLP showed 100 bands, of which 92 were polymorphic (92%). The fragments were scored as present (1) or absent (0), and those readings were entered in a computer file as a binary matrix (one for each marker). Three cluster analyses were performed to express–in the form of dendrograms–the relationships among the genotypes and the genetic variability detected. All DNA-based techniques used were able to amplify all of the genotypes. There were highly significant correlation coefficients between cophenetic matrices based on the genetic distance for the RAPD, ISSR, AFLP, and combined RAPD-ISSR-AFLP data (0.68, 0.78, 0.70, and 0.70, respectively). Two hypotheses were formulated to explain these results; both of them are in agreement with the results obtained using these three types of molecular markers. We conclude that when we study genotypes close related, the analysis of variability could require more than one DNA-based technique; in fact, the genetic variation present in different sources could interfere or combine with the more or less polymorphic ability, as our results showed for RAPD, ISSR and AFLP markers. Our results indicate that AFLP seemed to be the best-suited molecular assay for fingerprinting and assessing genetic relationship among genotypes of Dactylis glomerata.     

辣椒种质遗传多样性的RAPD和ISSR及其表型数据分析

用RAPDI、SSR分子标记及28个表型性状数据对辣椒属5个栽培种的13份材料进行了分析,结果表明:23条RAPD引物共扩增出209条带,平均每个引物扩增出9.09条,多态性位点比率为83.73%;16条ISSR引物共扩增出94条带,平均每个引物扩增出5.88条,多态性位点比率为79.79%.与RAPD相比,ISSR标记检测到的有效等位基因数(Ne)及Shannon多样性指数(I)、遗传离散度(Ht)和遗传分化系数(Gst)等遗传多样性参数都较大,多态性位点比例在亲缘关系较近的一年生辣椒(Capsicum annuum)种内较高,说明ISSR有更高的多态性检测效率,并且适合亲缘关系较近的种群间遗传多样性分析.基于RAPDI、SSR的聚类与基于表型数据的聚类之间存在极显著正相关,且都能将C.annuum与其它栽培种区分开来.     

ISSR and RAPD based evaluation of genetic fidelity and active ingredient analysis of regenerated plants of Picrorhiza kurroa

Genetic stability and phytochemical analysis of in vitro established plants of Picrorhiza kurroa Royle ex Benth, have been carried out. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) markers were used to assess the genetic fidelity of tissue culture products including three adventitious shoots from three calli and 6 months old tissue culture raised plants growing in green house condition with mother plant. Apparent genetic variation was detected in the five types of plant materials. The percentage of polymorphic bands in the RAPD and ISSR analysis were 16.25 and 14.54 %, respectively. The genetic similarity was calculated on the basis of RAPD and ISSR data among the five types of plant materials and were ranged from 0.5 to 1.0 (mean 0.75) and 0.47 to 1.0 (mean 0.73), respectively. The similarity coefficient by both RAPD and ISSR analysis revealed that differences between tissue culture raised plants and mother plant was not remarkable, but notable differences were observed among three adventitious shoots regenerated from three calli. The phytochemical analysis of tissue culture raised products showed higher secondary metabolite (picrotin and picrotoxinin) content as compare to mother plant. The information gained on genetic stability/variability will be valuable for the large scale propagation and secondary metabolite production of P. kurroa.     

Comparison of RAPD and ISSR markers for genetic diversity analysis among different endangered Mangifera indica genotypes of Indian Gir forest region

The genetic variability and relationships among 20 Mangifera indica genotypes representing 15 endangered and 5 cultivars, obtained from Indian Gir forest region, were analyzed using 10 random amplified polymorphic DNA (RAPD) and 21 inter simple sequence repeat (ISSR) markers. RAPD markers were more efficient than the ISSR assay with regards to polymorphism detection. Also, the average numbers of polymorphic loci per primer, average polymorphic information content (PIC) and primer index (PI) values were more for RAPD than for ISSR. But, total number of genotype specific marker loci, Nei’s genetic diversity (h), Shannon’s information index (I), total heterozygosity (Ht), average heterozygosity (Hs) and mean coefficient of gene differentiation (Gst) were more for ISSR as compared to RAPD markers. The regression test between the two Nei’s genetic diversity indexes showed low regression between RAPD and ISSR based similarities but maximum for RAPD and RAPD + ISSR based similarities. The pattern of clustering of genotypes within groups was not similar when RAPD and ISSR derived dendrogram were compared. Thus, both the markers were equally important for genetic diversity analysis in M. indica.     

Comparative assessment of ISSR, DAMD and SCoT markers for evaluation of genetic diversity and conservation of landrace chickpea (Cicer arietinum L.) genotypes collected from north-west of Iran

Morphological traits and three molecular markers techniques: start codon targeted (SCoT), inter-simple sequence repeat (ISSR) and directly amplified minisatellite DNA (DAMD) markers were compared for fingerprinting of 40 landraces chickpea genotypes collected from different geographical locations of north-west of Iran. Variance analysis of ten measured morphological traits showed significant differences existed between genotypes. Cluster analysis based on morphological traits, divided genotypes in three distinct clusters. Average polymorphism information content (PIC) for ISSR, DAMD and SCoT markers was 0.216, 0.232 and 0.232, respectively, and this revealed that SCoT markers were more informative, followed by ISSRs marker, than other markers for the assessment of diversity amongst genotypes. Cluster analysis for three different molecular types revealed that genotypes taken for the analysis can be divided in three and four distinct clusters. Accessions from same geographical regions mostly showed more genetic similarities than those from origins far isolated apart. These results suggest that efficiency of SCOT, DAMD and ISSR markers was relatively the same in fingerprinting of genotypes but SCOT and DAMD analysis are more effective in fingerprinting of chickpea genotypes. To our knowledge, this is the first detailed report of a comparison of performance among two targeted DNA region molecular markers (SCoT and DAMD) and the ISSR technique on a set of samples of chickpea. Overall, our results indicate that SCOT, ISSR and DAMD fingerprinting could be used to detect polymorphism for genotypes of chickpea.     

Multivariate analysis to determine the genetic distance among backcross papaya (Carica papaya) progenies

Morpho-agronomic and molecular (RAPD and ISSR markers) data were used to evaluate genetic distances between papaya backcross progenies in order to help identify agronomically superior genotypes. Thirty-two papaya progenies were evaluated based on 15 morpho-agronomic characteristics, 20 ISSR and 19 RAPD primers. Manhattan, Jaccard and Gower distances were used to estimate differences based on continuous and binary data and combined analyses, respectively. Except for production, there were significant differences in the continuous variables among the genotypes. The molecular analysis revealed 193 dominant markers (ISSR and RAPD), being 53 polymorphic loci. Among the various clusters that were generated, the one based on a combined analysis of morpho-agronomic and molecular data gave the highest cophenetic correlation (0.72) compared to individual analysis, consistently allocating the progenies into six groups. We found that the Gower algorithm was more coherent in the discrimination of the genotypes, demonstrating that a combination of molecular and agronomic data is valuable for studies of genetic dissimilarity in papaya.     

利用RAPD和ISSR分子标记分析怀地黄种质遗传多样性

用RAPD与ISSR技术对怀地黄的8个品种和2个脱毒品系进行了种质遗传多样性分析。分别从80条RAPD引物和44条ISSR引物中筛选出适合怀地黄种质分析的17条RAPD引物和10条ISSR引物,用于RAPD和ISSR分析。17条RAPD引物共扩增出177条带, 多态性位点数为109; 多态性位点比率为61.58%;平均多样性指数(I)为0.3135;每个位点的有效等位基因数（Ne）是1.3641; 10条ISSR引物共扩增出110条带. 多态性位点数为79; 多态性位点比率为71.58%;平均多样性指数(I)为0.3577;每个位点的有效等位基因数（Ne）是1.4037。 基于扩增条带数据库建立了各自的Jaccard遗传相关系数矩阵,构建了相似的分子树状图,将10个供试材料分为2类:一类群含组培85.5、大田85.5、组培9302、大田9302、金状元和金白6个材料;另一类群含北京1号、大红袍、地黄9104和野生地黄4个材料。两种分子标记的分析结果呈极显著正相关(r＝0.649)。结果表明,RAPD与ISSR标记适合于怀地黄种质遗传多样性分析,ISSR标记技术是一种多态性和重复性优于RAPD技术的实用技术。     

A Comparative Analysis of ISSR and RAPD Markers for Study of Genetic Diversity in Shisham (<Emphasis Type="Italic">Dalbergia sissoo</Emphasis>)

Shisham (Dalbergia sissoo) is one of the most preferred timber tree species of South Asia. Two DNA-based molecular marker techniques, intersimple sequence repeat (ISSR) and random amplified polymorphism DNA (RAPD), were compared to study the genetic diversity in this species. A total of 30 polymorphic primers (15 ISSR and 15 random) were used. Amplification of genomic DNA of 22 genotypes, using ISSR analysis, yielded 117 fragments, of which 64 were polymorphic. Number of amplified fragments with ISSR primers ranged from five to ten and varied in size from 180 to 1,900 bp. Percentage polymorphism ranged from 0 to 87.5. The 15 RAPD primers produced 144 bands across 22 genotypes, of which 84 were polymorphic. The number of amplified bands varied from five to 13, with size range from 180 to 2,400 bp. Percentage polymorphism ranged from 0 to 100, with an average of 58.3 across. RAPD markers were relatively more efficient than the ISSR assay. The mental test between two Jaccard’s similarity matrices gave r ≥ 0.90, showing very good fit correlation in between ISSR- and RAPD-based similarities. Clustering of isolates remained more or less the same in RAPD and combined data of RAPD and ISSR. The similarity coefficient ranged from 0.734 to 0.939, 0.563 to 0.946, and 0.648 to 0.920 with ISSR, RAPD, and combined dendrogram, respectively.     

Correspondence of ISSR and RAPD markers for comparative analysis of genetic diversity among different apricot genotypes from cold arid deserts of trans-Himalayas

The phylogenetic relationships of 36 locally grown Prunus armeniaca genotypes which are collected from nine sampling sites from two valleys viz. Nubra (9,600 ft) and Leh (11,500 ft) of trans-Himalayan region were analyzed using 31 PCR markers (20 RAPDs and 11 ISSRs). This is the first report of molecular genetic diversity studies in apricot from this region of the world. RAPD analysis yielded 139 fragments, of which 136 were polymorphic, with an average of 6.8 polymorphic fragments per primer. ISSR analysis produced 58 bands, of which 56 were polymorphic, with an average of 5.09 polymorphic fragments per primer. The primers based on (CT)n produced maximum number of bands (nine) while, (AT)n and many other motifs gave no amplification. RAPD markers were found more efficient with regards to polymorphism detection, as they detected 97.84 % as compared to 96.5 % for ISSR markers. Clustering of genotypes within groups was not similar when RAPD and ISSR derived dendrogram were compared, whereas the pattern of clustering of the genotypes remained more or less the same in RAPD and combined data of RAPD + ISSR. The results of PCA analysis were comparable to the cluster analysis. These analyses, allowed us to identify the groups corresponding to the two apricot collection sites.Key words: Prunus armeniaca, Apricot, Genetic Diversity, RAPD, ISSR, AMOVA     

R-ISSR marker as a useful tool for detection of new genomic loci in <Emphasis Type="Italic">Arthrocnemum macrostachyum</Emphasis>

Arthrocnemum macrostachyum, is a perennial halophytic shrub typical of Mediterranean salt marshes. The present study aims to investigate some combinations of inter simple sequence repeat (ISSR) and random amplified polymorphic DNA (RAPD) primers applied in real PCR. Thereby, the potential of R-ISSR markers to detect new genomic loci in 3 genotypes of A. macrostachyum grown in the Western coast of Syria was examined. Different combinations of RAPD and ISSR primers produced bands that were absent when single ISSR or RAPD primers were used. The results have demonstrated that ISSR primer (AG)₈TC gave more informative pattern when combined with different RAPD primers comparing to other tested primers. In contrast, the tested ISSR primer (GACA)₄ gave less informative pattern when used alone. These combinations were successfully applied in real PCR to detect new genomic variability in A. macrostachyum genotypes.     

Molecular characterization of <Emphasis Type="Italic">Vigna radiata</Emphasis> (L.) Wilczek genotypes based on nuclear ribosomal DNA and RAPD polymorphism

Mungbean germplasm characterization, evaluation and improvement are fundamentally based on morpho-agronomic traits. The lack of break-through in mungbean production has been due to non-availability of genetic variability for high yield potential. Forty-four genotypes of mungbean [Vigna radiata (L.)Wilczek] were subjected to random amplified polymorphic DNA (RAPD) analysis to assess the genetic diversity and relationships among the genotypes. Multilocus genotyping by twelve RAPD primers generated 166 markers and detected an average of intraspecific variation amounting to 82% polymorphism in banding patterns. Dendrogram obtained from cluster analysis delineated all the 44 genotypes into six clusters. Higher values of Nei’s gene diversity (h) and Shannon information index (i) and genetic distance analysis validate existence of wide genetic diversity among mungbean genotypes tested. Besides internal transcribed spacer (ITS) length variations, single nucleotide polymorphisms (SNPs) and insertions/deletions (INDELS) were detected at number of sites in nuclear rDNA region and the sequences of representatives of each sub-cluster and all distinct genotypes have been submitted to NCBI database and assigned Gen accession numbers HQ 148136-148147. Multiple sequence alignment revealed further lineages of distinct genotypes with main RAPD clusters. The measures of relative genetic distances among the genotypes of mungbean did not completely correlate the geographical places of their development. The homogeneous phenotypic markers proved insufficient in exhibiting genetic divergence among mungbean genotypes studied. RMG-62, RMG-976, and NDM-56 have been identified as potential source of parents for crop improvement. RAPD primers, OPA-9 and OPA-2 as polymorphic genetic markers and number of pods/plant and number of seeds/plant as dependable phenotypic markers have been identified for improving yield potentials. This genetic diversity will be of significance in developing intraspecific crosses in mungbean crop improvement programme.     

Analysis of genetic stability of in vitro propagated potato microtubers using DNA markers

The genetic stability of in vitro propagated potato microtubers was assessed using random amplified polymorphic DNA (RAPD), inter simple sequence repeat (ISSR), simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers. Microtubers were developed through in vitro from potato microplants using standardized protocols. The microtubers were conserved for 1 year under three different culture media and consequently microplants were regenerated for the DNA analyses. During the study, a total of 38 (10 RAPD, 11 ISSR, 12 SSR and 5 AFLP) primers produced a total of 407 (58 RAPD, 56 ISSR, 96 SSR and 197 AFLP) clear, distinct and reproducible amplicons. Cluster analysis revealed 100 % genetic similarity among the mother plant and its derivatives within the clusters by SSR, ISSR and RAPD analyses, whereas AFLP analysis revealed from 85 to 100 % genetic similarity. Dendrogram analysis based on the Jaccard’s coefficient classified the genotypes into five clusters (I–V), each cluster consisting of mother plant and its derivatives. Principal component analysis (PCA) also plotted mother plant and its genotypes of each cluster together. Based on our results, it is concluded that AFLP is the best method followed by SSR, ISSR and RAPD to detect genetic stability of in vitro conserved potato microtubers. The in vitro conservation medium (T2) is a safe method for conservation of potato microtubers to produce true-to-type plans.     

Isolation and HPLC assisted quantification of two iridoid glycoside compounds and molecular DNA fingerprinting in critically endangered medicinal Picrorhiza kurroa Royle ex Benth: implications for conservation

Picrorhiza kurroa is a medicinally important, high altitude perennial herb, endemic to the Himalayas. It possesses strong hepato-protective bioactivity that is contributed by two iridoid picroside compounds viz Picroside-I (P-I) and Picroside-II (P-II). Commercially, many P. kurroa based hepato-stimulatory Ayurvedic drug brands that use different proportions of P-I and P-II are available in the market. To identify genetically heterozygous and high yielding genotypes for multiplication, sustained use and conservation, it is essential to assess genetic and phytochemical diversity and understand the population structure of P. kurroa. In the present study, isolation and HPLC based quantification of picrosides P-I and P-II and molecular DNA fingerprinting using RAPD, AFLP and ISSR markers have been undertaken in 124 and 91 genotypes, respectively. The analyzed samples were collected from 10 natural P. kurroa Himalayan populations spread across four states (Jammu & Kashmir, Sikkim, Uttarakhand and Himachal Pradesh) of India. Genotypes used in this study covered around 1000 km geographical area of the total Indian Himalayan habitat range of P. kurroa. Significant quantitative variation ranging from 0.01 per cent to 4.15% for P-I, and from 0.01% to 3.18% in P-II picroside was observed in the analyzed samples. Three molecular DNA markers, RAPD (22 primers), ISSR (15 primers) and AFLP (07 primer combinations) also revealed a high level of genetic variation. The percentage polymorphism and effective number of alleles for RAPD, ISSR and AFLP analysis varied from 83.5%, 80.6% and 72.1%; 1.5722, 1.5787 and 1.5665, respectively. Further, the rate of gene flow (Nm) between populations was moderate for RAPD (0.8434), and AFLP (0.9882) and comparatively higher for ISSR (1.6093). Fst values were observed to be 0.56, 0.33, and 0.51 for RAPD, ISSR and AFLP markers, respectively. These values suggest that most of the observed genetic variation resided within populations. Neighbour joining (NJ), principal coordinate analysis (PCoA) and Bayesian based STRUCTURE grouped all the analyzed accessions into largely region-wise clusters and showed some inter-mixing between the populations, indicating the existence of distinct gene pools with limited gene flow/exchange. The present study has revealed a high level of genetic diversity in the analyzed populations. The analysis has resulted in identification of genetically diverse and high picrosides containing P. kurroa genotypes from Sainj, Dayara, Tungnath, Furkia, Parsuthach, Arampatri, Manvarsar, Kedarnath, Thangu and Temza in the Indian Himalayan region. The inferences generated in this study can be used to devise future resource management and conservation strategies in P. kurroa.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12298-021-00972-w.     

Comparison of RAPD and ISSR markers for assessment of genetic diversity among endangered rare Dalbergia oliveri (Fabaceae) genotypes in Vietnam

Dalbergia oliveri is a leguminous tree of the Fabaceae family. This species is popular and valuable in Vietnam and is currently listed on the Vietnam Red List and on the IUCN Red List as endangered. Two PCR techniques using RAPD and inter-simple sequence repeat (ISSR) markers were used to make a comparative analysis of genetic diversity in this species. Fifty-six polymorphic primers (29 RAPD and 27 ISSR) were used. The RAPD primers produced 63 bands across 35 genotypes, of which 24 were polymorphic. The number of amplified bands varied from one to four, with a size range from 250 to 1400 bp. The percentage polymorphism ranged from 0 to 75. Amplification of genomic DNA of the 35 genotypes, using ISSR analysis, yielded 104 fragments, of which 63 were polymorphic. The number of amplified fragments using ISSR primers ranged from one to nine and varied in size from 250 to 1500 bp. The percentage polymorphism ranged from 0 to 100. ISSR markers were relatively more efficient than RAPDs. The mental test between two Jaccard's similarity matrices gave r ≥0.802, showing good fit correlation between ISSRs and RAPDs. Clustering of isolates remained more or less the same for RAPDs compared to combined RAPD and ISSR data. The similarity coefficient ranged from 0.785 to 1.000, 0.698 to 0.956 and 0.752 to 0.964 with RAPD, ISSR, and the combined RAPD-ISSR dendrogram, respectively.     

Genetic variation,population structure and identification of yellow catfish, <Emphasis Type="Italic">Mystus nemurus</Emphasis> (C&;V) in Thailand using RAPD,ISSR and SCAR marker

Random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers were used to investigate the genetic structure of four subpopulations of Mystus nemurus in Thailand. The 7 RAPD and 7 ISSR primers were selected. Of 83 total RAPD fragments, 80 (96.39%) were polymorphic loci, and of 81 total ISSR fragments, 75 (92.59%) were polymorphic loci. Genetic variation and genetic differentiation obtained from RAPD fragments or ISSR fragments showed similar results. Percentage of polymorphic loci (%P), observed number of alleles, effective number of alleles, Nei’s gene diversity (H) and Shannon’s information index revealed moderate to high level of genetic variations within each M. nemurus subpopulation and overall population. High levels of genetic differentiations were received from pairwise unbiased genetic distance (D) and coefficient of differentiation. Mantel test between D or gene flow and geographical distance showed a low to moderate correlation. Analysis of molecular variance indicated that variations among subpopulations were higher than those within subpopulations. The UPGMA dendrograms, based on RAPD and ISSR, showing the genetic relationship among subpopulations are grouped into three clusters; Songkhla (SK) subpopulation was separated from the other subpopulations. The candidate species-specific and subpopulation-specific RAPD fragments were sequenced and used to design sequence-characterized amplified region primers which distinguished M. nemurus from other species and divided SK subpopulation from the other subpopulations. The markers used in this study should be useful for breeding programs and future aquacultural development of this species in Thailand.     

Evaluation of genetic diversity of <Emphasis Type="Italic">Clinacanthus nutans</Emphasis> (Acanthaceaea) using RAPD,ISSR and RAMP markers

Three polymerase chain reaction (PCR) techniques were compared to analyse the genetic diversity of Clinacanthus nutans eight populations in the northern region of Peninsular Malaysia. The PCR techniques were random amplified polymorphic deoxyribonucleic acids (RAPD), inter-simple sequence repeats (ISSR) and random amplified microsatellite polymorphisms (RAMP). Leaf genomic DNA was PCR amplified using 17 RAPD, 8 ISSR and 136 RAMP primers . However, only 10 RAPD primers, 5 ISSR primers and 37 RAMP primers produced reproducible bands. The results were evaluated for polymorphic information content (PIC), marker index (MI) and resolving power (RP). The RAMP marker was the most useful marker compared to RAPD and ISSR markers because it showed the highest average value of PIC (0.25), MI (11.36) and RP (2.86). The genetic diversity showed a high percentage of polymorphism at the species level compared to the population level. Furthermore, analysis of molecular variance revealed that the genetic diversity was higher within populations, as compared to among populations of C. nutans. From the results, the RAMP technique was recommended for the analysis of genetic diversity of C. nutans.     

A comparative analysis of genetic diversity in blackgram genotypes using RAPD and ISSR markers

Random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) markers were used to study the DNA polymorphism in elite blackgram genotypes. A total of 25 random and 16 ISSR primers were used. Amplification of genomic DNA of the 18 genotypes, using RAPD analysis, yielded 104 fragments that could be scored, of which 44 were polymorphic, with an average of 1.8 polymorphic fragments per primer. Number of amplified fragments with random primers ranged from two (OPA-13) to nine (OPK-4) and varied in size from 200 bp to 2,500 bp. Percentage polymorphism ranged from 16.6% (OPK-7) to a maximum of 66.6% (OPE-5, OPH-2, and OPK-8), with an average of 42.7%. The 16 ISSR primers used in the study produced 101 bands across 18 genotypes, of which 55 were polymorphic. The number of amplified bands varied from two (ISSR 858) to ten (ISSR 810), with a size range of 200–2,200 bp. The average numbers of bands per primer and polymorphic bands per primer were 6.3 and 3.4, respectively. Percentage polymorphism ranged from 25% (ISSR 885) to 100% (ISSR 858), with an average percentage polymorphism of 57.5% across all the genotypes. The 3-anchored primers based on poly(GA) and poly(AG) motifs produced high average polymorphisms of 54.98% and 58.32%, respectively. ISSR markers were more efficient than the RAPD assay, as they detected 57.4% polymorphic DNA markers in Vigna mungo as compared to 42.7% for RAPD markers. The Mantel test between the two Jaccards similarity matrices gave r =0.32, showing low correlation between RAPD- and ISSR-based similarities. Clustering of genotypes within groups was not similar when RAPD and ISSR derived dendrogram were compared, whereas the pattern of clustering of the genotypes remained more or less the same in ISSR and combined data of RAPD and ISSR.     

分子标记鉴定常山胡柚优良基因型的初步研究

本研究利用RAPD和ISSR分子标记对常山胡柚的优良基因型进行鉴定,并探讨常山胡柚的起源。从100个RAPD引物中筛选出12个多态性引物用于正式扩增,共得到117条DNA带,其中多态性DNA带64条,占扩增片段的54．7%;从105个ISSR引物中筛选出11个多态性引物用于正式扩增,共得到94条DNA带,其中多态性DNA带58条,占扩增片段的61．7%。RAPD和ISSR分析揭示了常山胡柚及其近缘种的一些特异性条带。ISSR共产生了15条特异条带,RAPD共产生12特异性条带。实验数据用AMOVA软件计算遗传距离,用NTSYS-pc软件构建UPGMA聚类树状图。结果显示,所有的基因型及不同种之间均能够彼此区分,分析得到的指纹图谱对常山胡柚种和基因型的鉴定具有潜在的应用价值,可用于优良基因型的鉴定。聚类分析结果显示常山胡柚和甜柚聚为一枝,确定了甜柚是杂交亲本之一,但是常山胡柚和柚的遗传距离较远,说明常山胡柚可能是甜橙、柚和柑桔属其他种的多重自然杂交的结果。     

Morphological and molecular genetic variations of oat genotypes grown in Kermanshah,Iran

Morphological traits and molecular markers are two common methods for genetic variation studies. Molecular markers, morphological traits methods and relationship between the two were used to study genetic variation among 43 oat genotypes and varieties. For this purpose, an augmented design was conducted in three replicates at 2008–2009 cropping season in the experimental field of Campus of Agriculture and Natural Resources of Razi University, Kermanshah, Iran. Four wild oat accessions (Avena sterilis) were added to evaluated genotypes in molecular experiment. Results showed a significant variation among genotypes for all morphological traits and they were classified based on this variation in four groups by WARD cluster analysis. In molecular experiment, 28 inter simple sequence repeat (ISSR) primers amplified 206 polymorph bands. Based on Jaccard similarity matrix, similarity among genotypes was varied from 0.23 to 0.66 and cluster analysis classified genotypes in seven groups by complete linkage method. The correlation between ISSR marker and morphological traits classifications was not significant. ISSR showed to be a helpful marker for genotype identity and separation as it put wild accessions in a group.