1- Ministry of Science
2- Agricultural Research, Education and Extension Organization, Gorgan, Iran
2- Agricultural Research, Education and Extension Organization, Gorgan, Iran
Abstract: (7 Views)
Contamination control of leaf and petiole explant of red maple (Acer Rubrum L.)
in vitro
Sheyda Amirian1, Seyed Mohammad Hosseini Nasr2, Hamid Jalilvand3and Akram Ahmadi 4
1. PhD student in Forest Biological Sciences, Sari University of Agricultural Sciences and Natural Resources, Faculty of Natural Resources, Sari, Iran (Corresponding author: sheyda.amirian3230@gmail.com)
2. Associate Professor, Sari University of Agricultural Sciences and Natural Resources, Faculty of Natural Resources, Department of Forestry, Sari, Iran.3. Professor, Sari University of Agricultural Sciences and Natural Resources, Faculty of Natural Resources, Department of Forestry, Sari, Iran.
4. Research Assistant Professor, Natural Resources Research Department, Golestan Province Agricultural and Natural Resources Research Center, Agricultural Research, Education and Extension Organization, Gorgan, Iran.
Abstract
Introduction: Standing red maple (Acer rubrum) native to the northeastern United States of America for various reasons, including the beauty of colored leaves, being suitable and widely used for urban green space design, resistance to cold, humidity and drought tolerance, and ecological characteristics and wide ability to Compatibility with different types of soil is one of the imported species of maple in Iran, which is difficult to reproduce through seeds and cuttings in high production, and due to the limitations created in terms of the season and the number of cuttings, favorable results in the scale It does not come with the top. Unfortunately, the success of laboratory cultivation methods is always hampered by microbial contamination, while contamination-free cultivation is a prerequisite for the success of laboratory-based plant biotechnology. Therefore, surface disinfection is a vital step in preparing healthy and living explants in tissue culture. Most surface contamination can be removed by disinfecting the surface with a suitable disinfectant. Since the studies conducted in the field of preparation of different sterile explants in the red maple genus are very few, this study was conducted with the aim of providing an optimal disinfection method for the micropropagation of the commercial species of red maple.
Materials and methods: First, the tested treatments were selected through library studies and research background checks. The required plant resources were obtained from seed organs, leaves and petioles of one to three-year-old young red maple seedlings from Mazandaran province, Shaheswar city, and the research procedures were carried out in the tissue culture laboratory of Golestan Agricultural and Natural Resources Research Center. Then, culture medium was prepared in six replicates of each treatment.
In the next step, the required explants were separated from the mother base, and after removing the damaged explants or those with signs of fungus, using a 500 cc container, the initial washing was done using Tiepol and 0.04% benomyl fungicide, and below The laminar hood was transferred and the disinfection process was continued there. The treatments were applied separately and the explants, after cutting into one centimeter pieces using tweezers and scalpel, were cultivated in jam glass containers with at least 20 cc of the intended culture medium and after being transferred to a light rack (12 hours of light) and 12 hours of darkness were placed in optimal environmental conditions with a temperature of 25 degrees Celsius and after seven days of cultivation, their contamination level was checked. Statistical analysis based on factorial experiment in the form of completely random basic design with three types of explants, two types of culture media and 13 treatments, each treatment was cultivated in five replications and three pieces of explants were cultivated in each replication. Finally, the data were analyzed using SPSS software.
Results: The findings of the present research showed that the sources of changes investigated in the experiment have a statistical difference with each other at the level of 0.01%. In this study, the diagnosis of fungal infections was done by morphological method and with the naked eye, which showed that the causative agent of fungal disease is Penicillium sp, Aspergillus sp, Cryptococcus sp, Mucur sp, Rhizopus sp, Fusarium sp, and Pythium sp. Based on the results of average comparisons, the rate of growth and spread of fungus in PDA culture medium with 83.2% showed more contamination than MS culture medium with 76.5%. In the comparison of the contamination rate of the used explants, it was found that the petiole explant with signs of contamination was (59.6%), less contamination than the leaf (8%) and the seed (96.2%). Finally, among all the applied treatments, the best treatment for decontamination of red maple explants for use in red maple micropropagation in Iran, Golestan province, (one drop of Teepol (two minutes) + benomyl fungicide (four grams per liter) ) (25 minutes) + 75% alcohol, (60 seconds) + 25% sodium hypochlorite, (15 minutes) + 0.1% mercury chloride (five minutes) was determined.
Conclusion: Maintaining disinfection conditions is a new definite prerequisite for the successful reproduction of species in laboratory conditions. Also, in the cultivation of ornamental and ornamental species such as red maple, which is aimed at commercial plant propagation, control of contamination and preparation of sterile rhizomes are of double importance due to the high volume of work. Because the smallest contamination in the explant quickly reproduces in the environment and causes a lot of financial damage, which is often not possible to compensate. In the present study, the use of disinfectants in the form of a combination of (Tipol (two minutes) + benomyl fungicide (4 g/liter) (25 minutes) + 75% alcohol, 60 seconds + sodium hypochlorite 25% (15 minutes) + chloride 0.1% mercury (five minutes) has a greater effect in controlling fungal infection of red maple seedlings. In general, in the present study, the use of any of these substances alone or in combination with another substance does not have a significant effect in decontamination of the red maple cultivation environment. The use of these materials is one of the most important methods of controlling bacterial and fungal contamination in red maple glass culture.
in vitro
Sheyda Amirian1, Seyed Mohammad Hosseini Nasr2, Hamid Jalilvand3and Akram Ahmadi 4
1. PhD student in Forest Biological Sciences, Sari University of Agricultural Sciences and Natural Resources, Faculty of Natural Resources, Sari, Iran (Corresponding author: sheyda.amirian3230@gmail.com)
2. Associate Professor, Sari University of Agricultural Sciences and Natural Resources, Faculty of Natural Resources, Department of Forestry, Sari, Iran.3. Professor, Sari University of Agricultural Sciences and Natural Resources, Faculty of Natural Resources, Department of Forestry, Sari, Iran.
4. Research Assistant Professor, Natural Resources Research Department, Golestan Province Agricultural and Natural Resources Research Center, Agricultural Research, Education and Extension Organization, Gorgan, Iran.
Abstract
Introduction: Standing red maple (Acer rubrum) native to the northeastern United States of America for various reasons, including the beauty of colored leaves, being suitable and widely used for urban green space design, resistance to cold, humidity and drought tolerance, and ecological characteristics and wide ability to Compatibility with different types of soil is one of the imported species of maple in Iran, which is difficult to reproduce through seeds and cuttings in high production, and due to the limitations created in terms of the season and the number of cuttings, favorable results in the scale It does not come with the top. Unfortunately, the success of laboratory cultivation methods is always hampered by microbial contamination, while contamination-free cultivation is a prerequisite for the success of laboratory-based plant biotechnology. Therefore, surface disinfection is a vital step in preparing healthy and living explants in tissue culture. Most surface contamination can be removed by disinfecting the surface with a suitable disinfectant. Since the studies conducted in the field of preparation of different sterile explants in the red maple genus are very few, this study was conducted with the aim of providing an optimal disinfection method for the micropropagation of the commercial species of red maple.
Materials and methods: First, the tested treatments were selected through library studies and research background checks. The required plant resources were obtained from seed organs, leaves and petioles of one to three-year-old young red maple seedlings from Mazandaran province, Shaheswar city, and the research procedures were carried out in the tissue culture laboratory of Golestan Agricultural and Natural Resources Research Center. Then, culture medium was prepared in six replicates of each treatment.
In the next step, the required explants were separated from the mother base, and after removing the damaged explants or those with signs of fungus, using a 500 cc container, the initial washing was done using Tiepol and 0.04% benomyl fungicide, and below The laminar hood was transferred and the disinfection process was continued there. The treatments were applied separately and the explants, after cutting into one centimeter pieces using tweezers and scalpel, were cultivated in jam glass containers with at least 20 cc of the intended culture medium and after being transferred to a light rack (12 hours of light) and 12 hours of darkness were placed in optimal environmental conditions with a temperature of 25 degrees Celsius and after seven days of cultivation, their contamination level was checked. Statistical analysis based on factorial experiment in the form of completely random basic design with three types of explants, two types of culture media and 13 treatments, each treatment was cultivated in five replications and three pieces of explants were cultivated in each replication. Finally, the data were analyzed using SPSS software.
Results: The findings of the present research showed that the sources of changes investigated in the experiment have a statistical difference with each other at the level of 0.01%. In this study, the diagnosis of fungal infections was done by morphological method and with the naked eye, which showed that the causative agent of fungal disease is Penicillium sp, Aspergillus sp, Cryptococcus sp, Mucur sp, Rhizopus sp, Fusarium sp, and Pythium sp. Based on the results of average comparisons, the rate of growth and spread of fungus in PDA culture medium with 83.2% showed more contamination than MS culture medium with 76.5%. In the comparison of the contamination rate of the used explants, it was found that the petiole explant with signs of contamination was (59.6%), less contamination than the leaf (8%) and the seed (96.2%). Finally, among all the applied treatments, the best treatment for decontamination of red maple explants for use in red maple micropropagation in Iran, Golestan province, (one drop of Teepol (two minutes) + benomyl fungicide (four grams per liter) ) (25 minutes) + 75% alcohol, (60 seconds) + 25% sodium hypochlorite, (15 minutes) + 0.1% mercury chloride (five minutes) was determined.
Conclusion: Maintaining disinfection conditions is a new definite prerequisite for the successful reproduction of species in laboratory conditions. Also, in the cultivation of ornamental and ornamental species such as red maple, which is aimed at commercial plant propagation, control of contamination and preparation of sterile rhizomes are of double importance due to the high volume of work. Because the smallest contamination in the explant quickly reproduces in the environment and causes a lot of financial damage, which is often not possible to compensate. In the present study, the use of disinfectants in the form of a combination of (Tipol (two minutes) + benomyl fungicide (4 g/liter) (25 minutes) + 75% alcohol, 60 seconds + sodium hypochlorite 25% (15 minutes) + chloride 0.1% mercury (five minutes) has a greater effect in controlling fungal infection of red maple seedlings. In general, in the present study, the use of any of these substances alone or in combination with another substance does not have a significant effect in decontamination of the red maple cultivation environment. The use of these materials is one of the most important methods of controlling bacterial and fungal contamination in red maple glass culture.
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