Shadi Attar; Gholamhossein Davarynejad; Leila Samiee; Mohammad Moghadam
Abstract
Introduction: Persian walnut (Juglans regia L.), belonging to the Juglandaceae family, has its natural origin in the mountainous regions of central Asia and especially northern forests of Iran. Most walnut genotypes are seedling and sexually reproduced. Conducting studies on the genetic structure of ...
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Introduction: Persian walnut (Juglans regia L.), belonging to the Juglandaceae family, has its natural origin in the mountainous regions of central Asia and especially northern forests of Iran. Most walnut genotypes are seedling and sexually reproduced. Conducting studies on the genetic structure of these genotypes to identify, select and maintain their genetic resources is important. Identifying and collecting local varieties of fruit trees is considered as the first step on the path of breeding programs and lack of information regarding plants genetic characteristics causes the breeding work to be done slowly. Various methods have been used for studying genetic diversity and determining the genetic relationship between European and Asian varieties of walnut and identifying commercial walnut varieties, among which we can mention: Morphologic indices, Alozyme, Isozym, RFLP, RAPD, AFLP and ISSR markers. ISSR molecular marker was used in order to investigate genetic diversity of some genotypes of Persian walnut (Juglans regia L.) in Mashhad orchards. .
Materials and methods: To begin with, about 56 walnut trees from 4 orchards in Mashhad (Esteghlal (1), Golestan (2), Alandasht (3) and Emam Reza (4)) were selected and tagged from 2014 to 2016. In the spring of 2014 with the beginning of trees growth and opening of leaves, a number of leaves from each genotype were collected. After DNA extraction, the quality of samples by agarose gel (1 percentage) and electrophoresis method and quantity of them via spectrophotometer device at 260 and 280 nm wavelengths were determined. First, 24 primers of ISSR marker were prepared and after initial evaluation on 5 random genotypes, 9 primers with high polymorphism and repeatability were selected for further investigation. For PCR reaction, Amplicon kit (code 180 301, made in Denmark) was used. Gel electrophoresis images of primers that produced polymorphic bands with suitable resolution were analyzed manually. After scoring the bands, in which 0 used for showing the absence of a band and 1 showing the presence of a band in each sample, 1 and 0 numbers were changed to matrix by using NTedit software. Genetic similarities of samples were calculated by using NTSYSpc software, SIMQUAL method and DICE similarity coefficient. Dendrogram by cluster analysis was drawn by using UPGMA method. Principle coordinate analysis (PCO) was performed using the NTSYS software and grouping samples were evaluated in a two-dimensional plot.
Results and discussion: Results showed that from 9 primers in total, 118 bands amplified were in 300 to 3000 base pairs, while 29 bands were polymorphic. The number of amplified fragment for each primer was different so that UBC 844 (14 bands) had the highest and UBC 890 (8 bands) had the lowest amplified bands. The average amplified fragments for each primer was 9.83. The percentage of polymorphic in various primers was different. Maximum polymorphism (80%) of the primers was observed in UBC830. In general, according to the percentage of polymorphic bands, low adjustment to any changes in the environment was indicated. This can be used as an indicator to determine the value of diversity and genetic erosion. In genotypes cluster analysis, clustering was performed based on Dice similarity coefficient and UPGMA method, and 10 clusters were formed. ISSR molecular marker somewhat revealed genetic diversity among walnuts genotypes, whereas the genetic diversity was lower than expected. In general, by reviewing the findings in other parts of the world about walnut genetic diversity and comparing them with the results of this study, despite existing high genetic diversity among walnuts in many areas, some reports of low genetic diversity among walnuts populations have been published and unfortunately in recent years these reports has made more attention. Based on the results of several studies reporting low genetic diversity among walnuts, the following factors can be effective in this problem: natural disasters, human impacts such as deforestation and selection and propagation of superior genotypes, and sometimes walnuts self - pollination. In this respect, there is concern that if this trend of decreasing genetic diversity in the walnut population continues, this invaluable crop will be in danger of extinction. So we should think about a remedy. Finally, this investigation can be used as a start for conducting more researches in the region to maintain and manage this valuable crop germplasm and maximize genetic diversity for performing breeding programs in the future.
shadan khorshidi; Gholamhossein Davarynejad; leila samiei; mohammad moghaddam
Abstract
Introduction: Most deciduous trees need low temperature to break flower bud dormancy. One of the most important abiotic stresses is low temperature which limits production of temperate fruits. Pear production has been considerably reduced in recent years. Important pear cultivars show different levels ...
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Introduction: Most deciduous trees need low temperature to break flower bud dormancy. One of the most important abiotic stresses is low temperature which limits production of temperate fruits. Pear production has been considerably reduced in recent years. Important pear cultivars show different levels of resistance to cold. Cold compatibility followed by resistance increase is controlled genetically and contains several mechanisms which lead to production of different metabolites such as: polypeptides, amino acids and sugars. The object of this research was to evaluate the frost resistance of different ‘Dare Gazi’ genotypes and other pear cultivars in Mashhad climate condition.
Materials and Methods: This study was conducted to investigate the frost resistance of 23 ‘Dare Gazi’ pear genotypes and nine other cultivars include: ‘William’s’, ‘Bell de june’, ‘Spadona’, ‘Koshia’, ‘Domkaj’, ‘Torsh’, ‘Sebri’ and ‘Tabrizi’. Plant material contained vegetative and reproductive buds of one-year-old shoot samples which were collected from 25-year old trees on March 2014, four days after winter cold (-6.6 °C) in three directions of trees and sent to the laboratory. Frost damages of vegetative and reproductive buds were investigated based on visual observations (%), electrolyte leakage (EC) and proline content. EC was measured with a Metrohm 644 digital conductivity meter and proline content was measured based on Bates et al. (1973) method, using acid ninhydrin. The experiment was performed on completely randomized experimental design with three replications. Statistical analysis was carried out using MSTAT-C and Excel software. Mean values were compared using the least significance difference test (LSD) at 1% levels. Cluster analysis was conducted by SPSS 16 program.
Results and Discussion: Highest EC of reproductive buds was observed in ‘Dare Gazi’ 10, 19, ‘Tabrizi’ and ‘Torsh’ whereas ‘Dare Gazi’ 8, 18 and ‘Bell de June’ had the lowest EC. Based on visual observations, the least percentage of damaged reproductive buds was observed in ‘Dare Gazi’ 22 while ‘William’s’ suffered from frost at the highest damage level (96%). Göndör and Tóth (1998) studied 13 pear cultivars by microscopic observations of flower buds and found that ‘Packham's Triumph’ was relatively resistant under Hungarian ecological conditions. Honty et al. (2008) reported that Kaiser was the most sensitive pear cultivar to low temperatures during endodormancy and ecodormancy. Khorshidi et al. (2014) described that pear reproductive buds of ‘Ghodumi’ were the most tolerant. Considering the vegetative buds, ‘Dare Gazi’ 19 had the highest EC (74.47 %) which was not significantly different from ‘Dare Gazi’ 10, 20 and 3 whereas the lowest one was found in ‘William’s’(24.75%). The highest percentage of healthy vegetative buds was found in ‘Dare Gazi’ 1 (50%) which did not show a significant difference with ‘Dare Gazi’ 12, 5 and 7. ‘Tabrizi’ was the most sensitive and had most damaged vegetative buds (95.99%). Khorshidi et al. (2014) observed the least damaged vegetative buds in ‘Dare Gazi’ and the most damaged buds in ‘Boheme’ and ‘Ghodumi’. Palonen and Buszard (1997) mentioned that hardiness of woody tissue of apples did not seem related to flower bud hardiness. The highest proline content of reproductive buds was found in ‘Dare Gazi’ 20 (21.28 µmol g-1FW) and the lowest content observed in ‘Dare Gazi’ 2 (0.1 µmol g-1 FW). Young (1977) described that increase in proline was not correlated well with relative cold hardiness of citrus rootstocks. Data did not show any significant correlation between EC and proline content. Barka and Audran (1997) studied grape buds and shoots and reached a high negative correlation between proline content and frosttolerance. Yelonsky (1979) indicated that accumulation of proline was not correlated to cold hardiness. The results of present experiment were in agreement with Duncan and Jack (1987) findings which reported that increase in proline was not always correspondent to cold resistance. Based on the cluster analysis, the genotypes laid in two distinct groups. First group included ‘Dare Gazi’1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 13, 14, 15, 17, 18, 19, 20, 23 and ‘Sebri’, ‘Domkaj’, ‘Bell de june’ and ‘Koshia’ and second group include ‘Dare Gazi’ 11,16, 21 and ‘William’s’, ‘Spadona’ and ‘Torsh’. Frost damage, EC and proline content were higher in the first group compared to second group.
Conclusion: Cosidering‘DareGazi’ genotypes, no correlation was found between proline content and frost damage rate. This shows that morphological differences among ‘Dare Gazi’ genotypes could be due to the existing of genetic variation of these genotypes or they are different clones of one cultivar.
