Iranian Walnut Research Articles

Study of Association Between Nut Traits and SCOT Molecular Marker in Some Old Iranian Walnut Trees

Study of Association Between Nut Traits and SCOT Molecular Marker in Some Old Iranian Walnut Trees

Composite core set construction and diversity analysis of Iranian walnut germplasm using molecular markers and phenotypic traits.

Iran is a center of origin and diversity for walnuts (Juglans regia L.) with very good potential for breeding purposes. The rich germplasm available, creates an opportunity for study and selection of the diverse walnut genotypes. In this study, the population structure of 104 Persian walnut accessions was assessed using AFLP markers in combination with phenotypic variability of 17 and 18 qualitative and quantitative traits respetively. The primers E-TG/M-CAG, with high values of number of polymorphic bands, polymorphic information content, marker index and Shannon's diversity index, were the most effective in detecting genetic variation within the walnut germplasm. Multivariate analysis of variance indicated 93.98% of the genetic variability was between individuals, while 6.32% of variation was among populations. A relatively new technique, an advanced maximization strategy with a heuristic approach, was deployed to develop the core collection. Initially, three independent core collections (CC1-CC3) were created using phenotypic data and molecular markers. The three core collections (CC1-CC3) were then merged to generate a composite core collection (CC4). The mean difference percentage, variance difference percentage, variable rate of coefficient of variance percentage, coincidence rate of range percentage, Shannon's diversity index, and Nei's gene diversity were employed for comparative analysis. The CC4 with 46 accessions represented the complete range of phenotypic and genetic variability. This study is the first report describing development of a core collection in walnut using molecular marker data in combination with phenotypic values. The construction of core collection could facilitate the work for identification of genetic determinants of trait variability and aid effective utilization of diversity caused by outcrossing, in walnut breeding programs.

DNA fingerprinting and genetic diversity analysis with SCoT markers of Persian walnut populations (Juglans regia L.) in Iran

Juglans regia L. is the most widely cultivated and economically important species of edible nuts. Iran is known as a center of origin of genetically rich walnut germplasm and expected to be found a large diversity within Iranian walnut populations. A detailed population genetic of local populations is useful for developing an optimal strategy for in situ conservation and can assist the breeders in crop improvement programmes. The main aim of the present study was genetic fingerprinting of 20 Persian walnut populations, including 3 wild and 17 cultivated populations by using SCoT markers. We also aimed to evaluate the discriminating power of SCoT markers in these populations. The correlation between genetic distance and geographical distance has also investigated. The results showed that SCoT marker has a good discrimination power and can differentiate the studied populations. This marker can be used to evaluate genetic diversity, identify genotypes and DNA finger printing of Persian walnut populations. The present study also revealed that there are at least two genetic groups in the studied Persian walnut populations and the geographical features of cultivated populations can be influenced grouping of them. The studied populations differed genetically and showed isolation by distance.

Genome-wide patterns of population structure and association mapping of nut-related traits in Persian walnut populations from Iran using the Axiom J. regia 700K SNP array

Persian plateau (including Iran) is considered as one of the primary centers of origin of walnut. Sampling walnut trees originating from this arena and exploiting the capabilities of next-generation sequencing (NGS) can provide new insights into the degree of genetic variation across the walnut genome. The present study aimed to explore the population structure and genomic variation of an Iranian collection of Persian walnut (Juglans regia L.) and identify loci underlying the variation in nut and kernel related traits using the new Axiom J. regia 700K SNP genotyping array. We genotyped a diversity panel including 95 walnut genotypes from eight Iranian provinces with a variety of climate zones. A majority of the SNPs (323,273, 53.03%) fell into the “Poly High Resolution” class of polymorphisms, which includes the highest quality variants. Genetic structure assessment, using several approaches, divided the Iranian walnut panel into four principal clusters, reflecting their geographic partitioning. We observed high genetic variation across all of the populations (HO = 0.34 and HE = 0.38). The overall level of genetic differentiation among populations was moderate (FST = 0.07). However, the Semnan population showed high divergence from the other Iranian populations (on average FST = 0.12), most likely due to its geographical isolation. Based on parentage analysis, the level of relatedness was very low among the Iranian walnuts examined, reflecting the geographical distance between the Iranian provinces considered in our study. Finally, we performed a genome-wide association study (GWAS), identifying 55 SNPs significantly associated with nut and kernel-related traits. In conclusion, by applying the novel Axiom J. regia 700K SNP array we uncovered new unexplored genetic diversity and identified significant marker-trait associations for nut-related traits in Persian walnut that will be useful for future breeding programs in Iran and other countries.

Gasification of Iranian walnut shell as a bio-renewable resource for hydrogen-rich gas production using supercritical water technology

Gasification in supercritical water (SCW) media is known as an efficient and promising technology for obtaining hydrogen-rich gas from dry and wet bio-renewable materials. Gasification of walnut shell as the main hard nutshell produced in Kurdistan Province of Iran was investigated using a stainless steel batch micro-reactor. Effects of reaction time in the range of 10–30 min, feed loading in the range of 0.06–0.18 g, and temperature in the range of 400–440 °C were investigated to determine the condition for maximum hydrogen yield. Furthermore, carbon gasification efficiency (CGE) and hydrogen gasification efficiency (HGE) were calculated according to the elemental analysis and the yields of gaseous products. Total gas yield and hydrogen yield were directly correlated with temperature. Steam reforming of walnut shell was favored at higher temperatures. Also, walnut shell loading was inversely correlated with total gas and hydrogen yields while production of methane was favored by higher loading of walnut shell. Furthermore, hydrogen yield increased first, when reaction time increased from 10 to 20 min, and then decreased. Maximum hydrogen yield of 4.63 mmol/g of walnut shell was obtained at 440 °C, walnut shell loading of 0.06 g and reaction time of 20 min.

Determination and speciation of Se(IV), Se(VI), and selenomethionine in iranian walnut samples by spectrophotometry after cloud point extraction

A sensitive and selective micelle-mediated speciation and preconcentration method has been developed for trace amount of inorganic and organic selenium prior to UV-Vis spectrophotometry determination. The method is based on cloud point extraction of Se(IV) ions with polyethylene glycol tert-octylphenyl ether (Triton X-114) in presence of 2,3-diaminonaphthalene (DAN) as a chelating agent. Se(IV) ions react with DAN in a surfactant solution yielding a hydrophobic complex at pH 3. The phase separation was accelerated by a 20 min centrifugation at 3500 rpm. The calibration graph obtained from Se(IV)-DAN complexes was linear in the concentration range of 5.0–400.0 ng/mL and detection limit was 9 ng/mL. The relative standard deviation was 1.6% in the determination of 80 ng/mL Se(IV). The proposed method was successfully applied to the speciation and determination of selenium species-Se(IV), Se(VI), selenomethionine—in walnut samples with satisfactory results.

GENETIC STRUCTURE OF SOME WILD WALNUT POPULATIONS IN IRAN

In this study, twelve wild Iranian walnut (Juglans regia L.) populations were screened using ten simple sequence repeat (SSRs) markers. Based on our results, a high level of genetic diversity was observed among and within populations with the number of alleles per locus ranging from 6 to 11, with a total of 83 alleles and 8.3 alleles and 4.9 effective alleles on average per loci. In individual populations, the observed heterozygosities ranged from 0.525 to 0.713 with an average of 0.659. The average genetic distance among populations equaled 0.452, ranging from 0.094 to 0.73. Cluster analysis, done by UPGMA, grouped all populations into four main groups according to their genetic distances. These results provide evidence of the high potential of Persian walnut populations of Iran for breeding programs.

BENZENE ADSORPTION ON ACTIVATED CARBON FROM WALNUT SHELL

The adsorption ability of activated carbon derived from walnut shell was investigated for removal of benzene existing in the rich amine stream of gas sweetening systems. Carbonaceous samples derived from Iranian walnut shell (WS) were prepared by the chemical activation method using ZnCl2 and H3PO4 as activating agents. Impregnation ratio, activation temperature, and retention time in the final temperature were studied as operating variables based on the Taguchi method. The obtained activated carbons were characterized by measuring their characteristics by scanning electron microscopy (SEM), FT-IR, BET, and iodine number techniques. According to the results, carbon activated by H3PO4 has higher BET surface area (1544 m2/g), while carbon activated by ZnCl2 has better ability for removal of benzene (about 61.22 mg/g in an initial concentration of 300 mg/L). Benzene uptake was found to increase with increase in initial concentration and contact time. Kinetic and equilibrium adsorption studies were investigated.

Reducing power and radical scavenging activities of phenolic extracts from Juglans regia hulls and shells

Iranian walnut shows genetic diversity and has a large distribution. Different species of these walnuts have many variations in genes resistant to dryness, diseases, pests and other factors. In order to compare the reducing power and radical scavenging activities of Iranian walnut hull and shell phenolic extracts, seven different cultivars (Duclouxiana, Fallax, Orientis, Mayette, Fernor, Mellanaise and Elit) were selected from Barandoz, Khaletabad, Jamalabad, Kahriz, Gaznagh, Ziva and Nazlo, West Azerbaijan province, Iran, in 2010. The fruits of these walnuts were collected, their hulls and shells were dried and ground, and then methanolic extracts were prepared from these hulls and shells. Total phenolic content was determined using the Folin-Ciocalteu (F–C) method. The extracts’ reducing power and scavenging capacity for radical nitrite, hydrogen peroxide and superoxide were evaluated. The results show that the antioxidant and antiradical activities of the walnut hull are higher than those of its shell phenolic extract among different cultivars of walnut. In addition, the reducing power of walnut hull and shell phenolic extract was positively correlated with the phenolic content and radical scavenging capacities of walnut hull and shell extracts were positively correlated with phenolic content and reducing power in different cultivars. Key words: Juglans regia, hull, shell, phenolic content, reducing power, radical scavenging.

Various Antioxidative Activities of Iranian Walnut (Juglans regia L.) Green Husks Hydroalcoholic Extract and its Effects on LDL Oxidation

Walnuts contain several antioxidant compounds which are thought to contribute to their biological properties. The present work investigated the various antioxidative activities of walnut (Juglans regia L.) green husks hydroalcoholic extract (W) and its effect on oxidation of LDL induced by CuSO4 in vitro by monitoring the formation of conjugated dienes and thiobarbituric acid reactive substances (TBARS). It was demonstrated that total antioxidant capacity of W was 307±30 nmol of ascorbic acid equivalents/g W. The W showed remarkable scavenging activity on 2,2-diphenyl-picrylhydrazyl (DPPH) (IC50 1819 μg/ml). Total phenolic content of W was 2705 ± 519 mg of gallic acid equivalents (GAE)/g W and total flavonoid content of W was 10.08 ± 0.18 mg of of quercetin equivalents/g W. Walnut (Juglans regia L.) green huske extract showed an increase rate of 87 %, 178 % and 202 % lag time at concentrations ranging from 0.2 μg/ml to 20 μg/ml, respectively. The W was able to inhibit LDL oxidation and decrease the resistance of LDL against oxidation. The inhibitory effects of W on LDL oxidation were dose-dependent at concentrations ranging from 0.2 μg/ml to 20 μg/ml. This study showed that W is an source easily accessible of natural antioxidants and prevents the oxidation of LDL in vitro and it may be suitable for use in food and pharmaceutical applications.

Genetic diversity of Cytospora chrysosperma isolates obtained from Iranian walnut trees using molecular markers

To study the genetic diversity of isolates of Cytospora chrysosperma , the main causal agent of Cytospora canker, we collected the infected samples from the major walnut cultivation areas in 12 provinces of Iran. 58 representative isolates out of a total of 147 isolates were selected according to their geographical origins to study their genetic diversity using random amplified polymorphic deoxyribonucleic acid (RAPD) marker. Ten of 20 random primers amplified reproducible polymorphic banding patterns. Cluster analysis of the data using Centroid method and Jaccard´s similarity coefficient, divided the isolates into six groups, showing a high genetic diversity among populations of C. chrysosperma . Although there was no correlation between geographical origins and the resulting groups of RAPD analysis, but the amount of observed polymorphism (94%) indicated the adequacy of the molecular marker RAPD, for studying genetic diversity in C. chrysosperma isolates. This is the first report on the study of genetic diversity of populations of C. chrysosperma using RAPD marker. Key words : Biodiversity, fungi, canker, deoxyribonucleic acid, random amplified polymorphic deoxyribonucleic acid, Iran.

Genetic diversity of Cytospora chrysosperma isolates obtained from Iranian walnut trees using molecular markers

To study the genetic diversity of isolates of Cytospora chrysosperma , the main causal agent of Cytospora canker, we collected the infected samples from the major walnut cultivation areas in 12 provinces of Iran. 58 representative isolates out of a total of 147 isolates were selected according to their geographical origins to study their genetic diversity using random amplified polymorphic deoxyribonucleic acid (RAPD) marker. Ten of 20 random primers amplified reproducible polymorphic banding patterns. Cluster analysis of the data using Centroid method and Jaccard´s similarity coefficient, divided the isolates into six groups, showing a high genetic diversity among populations of C. chrysosperma . Although there was no correlation between geographical origins and the resulting groups of RAPD analysis, but the amount of observed polymorphism (94%) indicated the adequacy of the molecular marker RAPD, for studying genetic diversity in C. chrysosperma isolates. This is the first report on the study of genetic diversity of populations of C. chrysosperma using RAPD marker. Key words : Biodiversity, fungi, canker, deoxyribonucleic acid, random amplified polymorphic deoxyribonucleic acid, Iran.

Analysis of genetic diversity among some Persian walnut genotypes ( Juglans regia L.) using morphological traits and SSRs markers

There is a high diversity among cultivated walnut trees in Iran due to its long time of seed propagation and vast area of cultivation. In this study some morphological characters as well as Simple Sequence Repeat (SSRs) markers were used to analyze the genetic diversity and relationships among 31 Iranian walnut genotypes along with four foreign cultivars. The nut weight ranged from 7.52 to 17.73 g, kernel weight from 4.00 to 9.83 g, and kernel percentage ratio from 38.78 to 67.05% among studied genotypes. In SSRs analysis, nine primer pairs were tested that produced 39 alleles ranging from 2 to 8, with a mean value of 5.10 allele per primer. The Iranian genotypes showed relatively high diversity both for their SSRs loci and morphological traits. Although the foreign cultivars (‘Serr’, ‘Vina’, ‘Franquette’ and ‘Lara’) clustered with each other, they also laid close and within the Iranian genotype. The results of the study provided us with valuable diversity among our genotypes which could be used for breeding studies and also showed the power of genetic markers for analysis and evaluation of this diversity.

Biological characterization of Iranian walnut (Juglans regia) leaves

In recent years, emphasis has been placed on the use of natural materials in the control and treatment of various infections, as some chemically synthesized drugs have undesirable side effects. In this study, the ethanol extract of Iranian walnut leaves was examined for antibacterial activities on Streptococcus mutans, Streptococcus salivarius, Streptococcus sanguinis, and Actinomyces viscosus using the microdilution method. Total phenols, flavonoids, and flavonols were also determined colorimetrically. The minimum inhibitory concentrations (MIC) for ethanolic extract ranged between 15.6 and 187.5 mg/mL and minimum bactericidal concentrations (MBC) ranged between 31.25 and 250 mg/mL. Total phenols were 410 ± 14.43 mg/g gallic acid equivalent, and total flavonols and flavonoids were 270 ± 22.33 and 330 ± 12.21 mg/g rutin equivalent, respectively. These findings show that Iranian walnut leaves have antibacterial effects on the 4 examined bacteria and may be a suitable alternative remedy for protection and treatment of dental plaque due to these microorganisms.

Removal of mercury from water by carbonaceous sorbents derived from walnut shell

The adsorption ability of a powdered activated carbon (PAC) derived from walnut shell was investigated in an attempt to produce more economic and effective sorbent for the control of Hg(II) ion from industrial liquid streams. Carbonaceous sorbents derived from Iranian walnut shell (WS) were prepared by chemical activation method using ZnCl 2 as an activating reagent. To the best of our knowledge, this adsorbent was not used before for removing mercury from water. Adsorption of Hg(II) from aqueous solutions was carried out under different experimental conditions by varying treatment time, metal ion concentration, adsorbent amount, pH and solution temperature. It was determined that Hg(II) adsorption follows both Langmuir and Freundlich isotherms as well as pseudo-second-order kinetics. It was also shown that Hg(II) uptake decreases with increasing pH of the solution. The proper choice of preparation conditions resulted in a microporous activated carbon with 0.45 g/cm 3 density, 737 mg/g iodine number and 780 m 2/g BET surface area. The monolayer sorption capacity of this optimum adsorbent was obtained as 151.5 mg/g.