نشریه تحقیقات ژنتیک و اصلاح گیاهان مرتعی و جنگلی ایران
سال بیستم شماره 1 (پیاپی 39، بهار 1391)

Carpinus Genus is an important element in the Hyrcanian forest. There are some ambiguities and disagreements among botanist about the genus. In the current research, useful molecular markers were used for revision of the genus. We used DNA barcode nrDNA ITS and trnH-psbA and compared the performance of the two markers. CTAB method was used in order to extracting of leaves genomic DNA. PCR product sequencing and alignment and phylogenetic analyses of the dataset were conducted according to maximum parsimony (MP) method by using PAUP* A software. Results indicated that nrDNA ITS is more efficient than trnH-psbA for separation of the species. The surveyed results show that different species of the genus are placed in two separate claudes. Before that C. Betulus form Arasbaran was assumed same and similar to C. betulus from Hyrcanian forest but the current finding showed that it was completely isolated and placed in the separate claude. Furthermore, Carpinus orientalis and Carpinus schuschaensis are Synonymous. Finally, we proposed that C. betulus from Arasbaran should be considered by botanists.

Hypericum perforatum L. is a medicinal plant abounding with secondary metabolites which have clinically proven anti-inflammatory, antiviral, antifungal, and antidepressant activities. In this study, the genetic diversity of 29 accessions from eight populations collected from eight provinces (Azerbaijan, Gilan, Tehran, Semnan, Kurdistan, Lorestan, Golestan and Ardabil) were assessed using Inter Simple Sequence Repeat (ISSR) markers. Twelve primers containing different simple sequence repeats (microsatellite) were used. The primers produced between 11 to 26 bands. Totally the markers produced 221 amplification products, out of which 196 bands were polymorphic. The results showed that polymorphism information content (PIC) of primers was 0.37. Primers 841Y-UBC and UBC-807 had the highest PIC (0.41). Cluster analysis based on Jacquard’s similarity coefficient using Unweight Pair Group Method with Arithmetic mean (UPGMA) indicated wide range of diversity across the studied accessions. The highest genetic distance was observed between 23335 (Ardabil) with 17982 and 14206 (both of West Azerbaijan) accessions and between 133337 and 17982 accessions (Gilan and Azerbaijan) revealing closer genetic relationship. The least genetic distance was observed between 13180 and 13347 from Gilan. Molecular variation within and among the populations (within and among provinces) were estimated 78% and 22% of total variance, respectively. The distribution reflects high genetic diversity between the accessions in each geographical area. The results revealed that ISSR markers could be efficiently used for genetic differentiation of the St. John’s Wort accessions.

Moringa peregrina is the only species of the Moringa genus, which is growing naturally, in Southeast part of Iran. The species has industrial and medicinal values as well as environmental importance, which is suffering genetic erosion in the nature. Seeds were sampled on single plants of six plant populations in the habitats of the species in the country. The seeds were sown and several early growth stage characteristics were recorded on the seedlings based on a statistical design. Since the single trees of each plant population were not the same also the progenies of each single mother plants were not the same, the collected data were analyzed based on a nested model. Results of the analysis revealed that the plant populations were not significantly different based on the studied seedling characteristics and the majority of the differences belonged to within populations rather than between populations. Using expected mean squares, the results of analysis of variance were divided to their components. The analysis also showed that trees within population component of variance were stronger than the other components and the population component was not remarkable. Although the growth characteristics of the tree species would be more visible on higher ages of the trees, but the negligible genetic variation of the characteristics on the valuable species may be due to genetic erosion.

In order to evaluate phylogenetic relationships among some Vicia species, subspecies and ecotypes from Iran, 17 and 14 entries were studied using seed storage proteins electrophoresis and RAPDs markers, respectively. Total seed storage proteins were extracted from the collected seeds and DNA was extracted from leaf samples of seedlings. Nei and Shanon genetic diversity indices didnt show considerable difference between RAPD and storage protein data. While the PIC and MI indices for protein markers were greater than those of RAPD markers. In the phylogeny tree based on protein markers, the subspecies and ecotypes of sativa species were classified in the same group. In RAPD analysis, the subspecies and ecotypes were classified in a larger group along with other species. Ecotypes of V. panonica and V. cracca were classified in another group. Constructing the phylogenetic tree using incorporated RAPD and protein markers produced better results than separated ones. Also, classifying the entries using PCoA method was in correspondence to the classifying by phylogenetic trees. The distance matrices from RAPD and protein data were compared using Mantel test and were not observed any significant correlation between the two data sets.

Jujube (Ziziphus jujube Mill.) is one of the important medicinal plants belong to Rhamnaceae family. Jujube containes mucilage, vitamins, pectin, alkaloids, phenol compounds, and fatty acids. Therefore, it is important in the pharmaceutical industries. In addition to medicinal uses, the species has ornamental use as well. It has also been widely used in preventing soil erosion and desertification. This research was performed to evaluate genetic diversity of Iranian jujube genotypes using molecular AFLP marker. Twenty nine Jujube genotypes were selected from different parts of the country. Their molecular characteristics were assessed by 15 AFLP primers. Similarity matrix was generated using the Jaccard coefficient. Cluster analysis was carried out by UPGMA algorithm and Principle Coordinate Analysis. Twelve primers showed polymorphism. The primers produced 689 bands of which 44 bands showed polymorphism. Based on the dendrogram, the genotypes were divided to eight groups at 0.75 similarities. AFLP marker precisely separated the Qom, Isfahan and Khorasan groups. This resolution was very precise, so that Khorasan group was absolutely pure and separated from other groups. Cluster analysis result was confirmed by principal components analysis. Qom and Isfahan alocated in the same group. Therefore, the two genotypes may be considered as a common origin with high confidence. High efficiency of the applied marker in separation of genotypes was significant.

In order to investigate variation within and among Quercus castaneifolia populations, seed source effects and their parent trees on several seedling characteristics, seeds were collected from 34 mother plants originated from the habitats named Chamestan, Lavich (located in Noor), Koohsar-Kandeh and Khorram-Chamaz (located in Behshahr) (125, 450, 150 and 630 m above sea level, respectively). The seeds were sown at Pasand research station (26 m above sea level). At the end of the first year growing season, seed origin effects were investigated on survival, collar diameter, seedling height, leaf length, leaf width, leaf shape and leaf area. The strongest and weakest seedlings on the different origins were also specified based on the studied characteristics. Results indicated that the variations among the populations were statistically significant. The seedlings originated from the mother plants of Chamestan and Khorram-Chamaz were in better conditions than the seedlings of other locations. Moreover, difference between the mother plants of each location was also significant at 1% level of probability, based on the most of the recorded characters that indicated suitable variation within all populations. PCA analysis reveals that collar diameter, seedling height, leaf length, leaf width, and leaf area described 70% of the variance. Cluster analysis classified the genotypes into six groups.

Fars province is one of the natural habitats of Pyrus glabra Boiss. in Iran. Fruit harvesting, deforestation, expansion of agriculture and overgrazing during recent years has threatened the wild pear forests and imposed detrimental effects on their natural regeneration. Therefore, investigation on propagation of P. glabra for afforestation is needed. This research was carried out to study the effects of different growth regulators, nutrient media, gelling agents, and carbohydrate sources on shoot multiplication of P. glabra. Shoot tips and nodal segments of In Vitro propagated shoots originated from seeds and current season growth shoots of selected trees were used as explants. Two concentrations of 0.8 or 1.0 mgl-1 BA and 0.01 mgl-1 IBA were optimum for shoot multiplication. Addition of GA3 to nutrient medium had negative significant effects on shoot fresh weight of explants. Main shoot length of explants cultured on Murashige and Skoog (MS) medium was significantly longer than those cultured on woody plant medium (WPM) and Driver Kuniyuki walnut (DKW) (half strength macronutrients) medium. Quality and morphology of shoots produced on DKW medium were better than other nutrient media. Explants cultured on media containing sucrose or glucose did not show any significant differences for measured growth indices, but both of them were significantly better than those cultured on the media containing fructose. Media solidified with Phytagel were significantly better than those solidified with Difco Bacto agar for shoot fresh weight. The quality of shoots produced on media solidified with Phytagel was better than those on media solidified with Difco Bacto agar.

Thymus daenensis Celak. is an endemic plant in Iran and In vitro culture is useful for its propagation. Hyperhydricity syndrome is a common problem in In vitro culture causing abnormalities in growth and development of in vitro plants. In this investigation we studied the effect of benzyladenine and salicylic acid on DNA methylation of hyperhydric shoots. For this purpose, seeds were cultured into glass jar having benzyladenine and salicylic acid. Active oxygen and antioxidant enzymes analyses were done. Proline and H2O2 levels were high in BA treated-plantlets and also in plantlets growing without hormone but SA that reversed hyperhydric shoots to normal status, reduced the content of prolin, H2O2 and antioxidant enzymes. No statistical differences were observed in organelles DNA methylation, but 5 µM SA decreased the nuclear DNA methylation. Defense activator SA may have triggered genes engaged in stress response with an increase of proteins. Probably the decrease of methylation under SA treatment may be related with the reversion of hyperhydricity.

Regarding cross-pollination and a wide range of genetic variation in Ziziphus spina christti (L.) populations, vegetative propagation through tissue culture techniques is necessary to reproduce the elite single plants. In this study effects of different levels of growth regulators on callogenesis and regeneration capabilities of callus of leaf explants of the species were evaluated. Leaf explants of aseptic seedlings were cultured on MS medium containing 2, 4-D, NAA, TDZ and BAP levels (0.5, 1 and 2 mg/l). MS medium with different levels of BAP, TDZ and Kin (1, 3 and 5 mg / liter) was used for regeneration. Rooting process was done on 1/2MS medium with different levels of IBA and NAA levels (0.5, 1 and 2 mg /l). Results showed that the medium containing 2, 4-D and TDZ at concentrations of 0.5 and 1 mg/l (x=10) had the highest rate of callus induction. Kin treatments (5 mg/l) and TDZ (1 mg/l) with averages of 8.84 and 8.95 had the highest rate of shoot production, but the shoot produced by TDZ treatments had an unusual shape, whereas, Kin treatments, produced normal shoots. Average rooting on IBA was higher than NAA treatments on all concentrations. One mg/l IBA showed the highest rate of rooting capability compared to the other treatments.

Ducrosia anethifolia (DC) Boiss as a medicinal plant is a biennial herb of the Apiaceae family. Its origin is Iran and possessed n=11 (2n = 2x = 22) chromosome number forming mainly both rod and ring bivalents with some quadrivalents and univalents. A variety of abnormal chromosomal behaviors including chromosomal stickiness, bridge, laggards, micronucleus and precocious chromosome migration were observed, albeit with low frequency. Meiotic index was estimated. Tetrads with 4 equal-sized cells were considered normal and any deviation as abnormal. Pollen fertility was estimated by stainability. Few meiotic abnormalities were observed; as a consequence, high pollen fertility was concluded. Well-filled pollen grains with stained nuclei were regarded as fertile while shriveled, smaller microspores and unstained pollen were counted as sterile. This study reveals that meiosis in the species is normal, with high meiotic index and regular bivalent formation in most of the studied cells. Furthermore, the pollen stainability was 97% in this case.

Capparis decidua is a desert plant resistant to longtime drought conditions, special to southern part of Iran (Hormozgan, Bushehr and Sistan and Baluchistan Provinces). Due to lack of seed germination, asexual regeneration of intact trees of the species by tissue culture may save genetic resources of the valuable species. This research was performed in order to use in vitro culture and proliferation of seedling and mature trees to regenerate Capparis decidua. Mature seeds were collected from selected trees in natural habitat of the species located in Sirik forests of Hormozgan Province during June. Embryos were aseptically sterilized and transferred in different mediums (MS N/2, MCM and Magenta GA7 vessels with vermiculite, peat and perlite in 1:1:1 ratio, irrigated with and without MS medium). After 3 weeks, 74% of the embryos produced seedlings. Shoot tips of the seedlings and mature trees, were sterilized and cultured on MCM and MS mediums. At shooting stage, MCM containing 0.3 mgl-1 BA, 0.5 mgl-1 2ip, 0.1 mgl-1 IBA and 0.5 mgl-1 GA3 was the best medium. The highest rate of rooting (89%) was achieved on MS (N/2) containing 0.5 mg/l IBA, in Magenta vessels containing sterile vermiculate and peat. The plantlets were successfully transferred to the soil at Natural Resource Research of Jask.

Mountain Rye (Secale montanum Guss.) is a perennial grass, and attractive for grazing and forage production on pastures, with average precipitation of 300 mm per year. Seed shattering is a major constrain for grain yield loss for growers of the species. The objectives of the research were to study genetic diversity and introduce factors affecting seed shattering for detecting seed shattering resistant plant populations. The experiment was conducted on eight populations at experimental field of Agricultural and Natural Resources Research Center of Khorasan Razavi, Iran, during 2006 and 2007. The measurements were done on seed maturity and shattering at seed setting stage by 10-days intervals for five times and the spike characteristics, including length, weight, internode length, and thickness of the rachilla, number of floret, seed length and width, and 1000 seed weight. Results showed that seeds were fully matured by late June and seed shading started after June from top of inflorescence to bottom from late June for sensitive populations to mid-September for seed shattering resistant types. There was genetic variation for shattering resistance with low frequency within the populations. Therefore, populations were divided into two groups including seed shattering and non-shattering by t-test (P<0.01). Also, comparison of the spike characteristics revealed that the most important reducing factors of seed shattering were spike weight, spike length, their ratio, seed weight on spike, and spike internodes length with increments of 75, 19, 31, 31, and 5.1%, respectively, comparing to the seed shattering populations. The results suggest that seed shattering in mountain ray was influenced by genetic factors and native seed shattering resistant populations can be found in the country.

Iran is one of the main centers for genetic diversity of sainfoin (Onobrychis Mill.). A total of 198 landraces of sainfoin (O. vicifolia Scop. Syn O. sativa Lam.) genetic resources, collected from all over the country were subjected in the study. The materials were sown in an experimental field and evaluated based on their agro-morphological traits. A high degree of genetic diversity for most of the agro-morphological traits was observed. Days to flowering revealed a diversity of 40 days. Therefore, the germplasm can be used as suitable materials for improving different classes of seed ripening. There were suitable genetic diversity for susceptibility to leaf powdery mildew and pod shattering traits. Result of regression analysis implied the stem color as a character for fast screening on high yield forage selection. According to the result of cluster analysis based on K-means and factor analysis, landraces were grouped into 3 distinct clusters. Clusters relations and main traits in each cluster have been disscused. A remarkable collection of sainfoin germplasm was evaluated; therefore, it can provide suitable materials for future sainfoin breeding programs.

Chelcheragh lily (Lilium ledebourii) is a wild endangered species which has been registered as a national heritage in Iran. As an ornamental plant, it has also the potential to be supplied to national and global flower markets. In this study, the genetic diversity of different populations of the species was carried out based on electrophoresis of seed storage protein and the comparison of morphological characteristics in five plant populations of the species in three provinces of Gilan, Mazandaran and Ardebil. Based on 13 morphological characteristics, plants from Damash (Gilan province) had the highest average of the most of the studied traits while the two regions of Dorfak and kelardasht (Gilan and Mazandaran provinces, respectively) were very similar and in a lower rank. Seeds of five populations of the species were collected and SDS-PAGE electrophoresis was performed. Most of the bands were in the range of low molecular weight proteins (18.4, 35 and 45 KDa) and the lowest number of bands was observed in the moderate molecular weights. Presence or absence of the bands on the electrophoresis profile were determined and scored. Using cluster analysis, the populations were classified. The results divided the samples into two categories. Dorfak and Kelardasht plant populations were allocated in the same cluster while Ardabil, Damash and Vazrood plant populations were allocated in the other clusters. Minimum distance between the populations of Ardebil and Damash likely indicates common genetic origin. Dorfak population and Kelardasht had the lowest gap. Maximum distance was observed between Damash and Kelardasht populations suggesting that the differences in the genes encoding seed storage proteins probably indicates the development of the two populations towards independent species.