Optimization of the conditions for production of synthetic seeds by encapsulation of axillary buds derived from minituber sprouts in potato (Solanum tuberosum) (original) (raw)
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PRODUCTION OF VIRUS-FREE POTATO SYNTHETIC SEEDS AND SEEDLINGS USING TISSUE CULTURE TECHNIQUES
Thesis
ABSTRACT This study comprised two parts. In the first part, Alfalfa mosaic virus (AMV) was isolated from naturally infected potato plants grown in Bader Center, Behera Governorate, Egypt. The isolated virus was biologically purified by single local lesions developed on Ch. amaranticolor leaves and propagated in potato cv. Daimond. The isolated virus was identified and its identity was confirmed serologically and molecularly. The virus was identified as such on the bases of host range and symptomatology, modes of transmission, serological reaction, molecular biology and ultrastructure changes in infected potato leaf cells. In the second part, three cultivars of potato (Solanum tuberosum L.) Ditta, Santana and Daimond cvs. were used to compare their responses to callus induction and embryonic callus criteria. The data of induction stage revealed that the Ditta cv. achieved superior embryonic callus induction 79.52% and it was investigated to somatic embryogensis. Inter simple sequence repeats (ISSR) marker were applied by seven ISSR primers successfully and showed 48 polymorphic bands out of a total of 127 bands with 81.1% polymorphism which can be considered as useful markers for study the effect of the treatments used. Histological and morphological means via handling section, de-waxing and staining examined by light microscope to discriminate the progression of specific stages and general definition of SE, evaluated potato AMV-free production from infected potato cv. Ditta via two techniques: chemotherapy using saliycilic, thioracile, 8-azaguanin, cuomarin and kinetin; also embryonic calli as somatic embryogenesis pathway. Serological tests by Enzyme-linked immunosorbent assay (ELISA) was used to evaluate the effectiveness of the techniques. Next step, standardized a suitable technique for nutrient encaplsulation of potato nodal segments (unipolar synthetic seeds) and somatic embryo (bipolar synthetic seeds) as synthetic seeds by using different concentrations of sodium alginate at 2%, 3% and 4% and all of them were complexed by Ca-salts either Ca (Na3)2 or Na Cl2 in the presence of distilled water, 1/4 or 1/2 MS medium, sorbitol or ABA as artificial endosperm to evaluate of it conversion and vitality after storage under conditions: temperature (21±1 Cº and 4Cº) and containers (empty and with 1/2 MS solid medium in 10ml). Algaination degree and cracking degree of capsule and survival percentage were numder-taken. After storage under different temperature and time, the following data were recorded, extended of regeneration and variability of nodes and SE were examined. The presence of capsules in the lab was influenced by the presence of capsule contents. The absence of salts of Murashige and Skoog in the capsule was better. The use of these capsules decreased the conversion rate. When sorbitol was added, it retained its ability to shift and also increased its duration storage. While, absisic doses not gave any conversion rate. All of this is done when incubating at a temperature of 4 ° C. The synthetic seeds were cultivated in a conversion medium. Cultivation of all survival converted synthetic seeds and adapted resulted in successive survival and vigor plantlets (70%).
Micropropagation of Potato Solanum tuberosum L Micropropagation of Potato Solanum tuberosum L
The effect of cytokinins and combination of cytokinins and auxins on in vitro microtuber formation and growth of two potato cultivars of potato (Solanum tuberosum L.) were evaluated. In the present study sprouts and nodal explants of potato, cultivars Agrija and Andrea, were cultured on MS medium, supplemented with different hormonal combinations. For sprouts as an initial explants were used MS + 4 mg/l KIN and MS + 2 mg/l BAP, and for nodal explants were used MS + 4mg/l KIN + 1mg/l IAA and MS + 2 mg/l BAP+1 mg/l NAA. For rapid sprouting clean popato tubers were in vivo treated with 2 ppm GA 3 . Between the two different explants (nodal segment and sprout) nodal cutting showed the better microtuber formation. The cultivar Agrija showed greater ability for in vitro propagation, with 2.14 tubers per shoot and 13.33% microtuber formation.
Growth of potato axillary bud cultures in vitro
Pak. J. Agrie. Sci, 1996
III vitro shoot cultures of two commercial cultivars of Solanum tuberosum, Desiree and Maris Piper, and of two wild species S. conunersonii and S. acaule, were established. Growth of axillary buds was assessed on two different media. Espinoza medium proved better fill'the ...
Micropropagation of Potato Solanum tuberosum L.
2012
The effect of cytokinins and combination of cytokinins and auxins on in vitro microtuber formation and growth of two potato cultivars of potato (Solanum tuberosum L.) were evaluated. In the present study sprouts and nodal explants of potato, cultivars Agrija and Andrea, were cultured on MS medium, supplemented with different hormonal combinations.
Stem cuttings from potatoes are frequently used in multiplication program for disease-free seeds. In vitro plantlets with stem cutting technique of three cultivars, i.e., 'Diamant', 'Lady Balfour' and 'Red Baron' were used as a good source material to develop tissue culture protocol of minitubers seed production in the greenhouse. The first protocol (traditional) takes eight months to get microtubers (G 0), including meristem tip culture, four subcultures, in vitro microtuberization, then seven months to produce minitubers (G 1), including storage of microtubers, growth and harvesting, and eight months to produce the minitubers (G 2) with total equals 23 months. The second protocol (modified) takes ten months to get micro-minitubers (G 0) through four in vitro subcultures and the plantlets were acclimatized and grown tell harvesting, and eight months to produce the minitubers (G 1) with total equal of 18 months. The third protocol (new procedure) designed to magnification of the productivity of potato minitubers seeds by increasing the number and reducing the time of its production during 2014-2015 seasons. This protocol reduced the subcultures by two times then were acclimatized like the second protocol to produce micro-minitubers (G 0) in eight months, then eight months to produce (G 1) through a procedure using stem cuttings technique induced to grow at the leaf nodes by removal the apical growing point, with a total of 16 months. The in vitro plantlets with stem cutting material source yielded significantly better seed potato micro-minitubers number/plant (217) than traditional (18) and modified protocols (8). Among the cultivars, 'Diamant' gave significantly a large minitubers number (123) per plant as compared to the other cultivars. This indicates that in vitro plantlets with stem cutting technique have potential to give a viable material for seed potato minitubers production under greenhouses. In addition, this production system shortens the time needed to get the number of minitubers with seven months than the traditional protocol. Upon completion of the program via three ways to the second generation (G 2) of minitubers seed, the production rate of the new procedure gave 48877 minitubers compared with 71 with the modified and 133 minitubers with the traditional protocols.
Alaska, Safran and Spunta are the most important varieties of potatoes used by Tunisian farmers. This study was carried out in four steps. First, study on regeneration of tissue culture protocol was studied using buds as an explant for initiation of culture in MS media supplemented with four different concentrations of an auxin : indole butyric acid (IBA). Growth proliferation showed that optimum regeneration rate was obtained with 0,5 mg/l of IBA. The regenerated plants were cultured using nodal cuttings as explants for further multiplication. In vitro tuberization involving a combination benzyladenine (BA) and paclobutrazol (PBZ) gave good tuberization rate. Yet, liquid media containing 5 mg/l of BA was optimal to produce microtubers for all cultivars. Microtubers were transplanted in the soil, cultured in glasshouse to produce minitubers, they produces,-7,2 healthy minitubers/plant. Microtubers cultured in medium containing sucrose (80 g/l) gave best number of minitubers/plant.
Journal of Mathematical and Fundamental Sciences, 2014
Somatic embryogenesis can be used as an alternative method to obtain high-quality potato seedlings. This research was conducted in order to evaluate: the potential of different types of explants to produce somatic embryos; combinations of 2,4-D and BAP for somatic embryo induction; and the ability of BAP to act as growth regulator for the somatic embryo maturation process. Young leaf and stem nodal sections were cultured in MS media supplemented with 2,4-D and BAP. The developing calli were then transferred into media containing several concentrations of 2,4-D and BAP. The embryogenic calli were then transferred into media containing 1-5 μM BAP. The results show that MS media containing 2,5 µM 2,4-D + 5 µM BAP and µM 2,4-D + 0.5 µM BAP were the most suitable for inducing somatic embryos from leaf explants and stem nodal section explants, respectively. The somatic embryos were well-developed in the MS media supplemented with 5 µM BAP. The stem nodal section explants were able to produce a higher percentage of globular, heart and torpedo stage embryos compared with the leaf explants. Based on this research, stem nodal section explants have a high potential for producing somatic embryos induced in medium containing 1 μM 2,4-D + 0.5 μM BAP.
Effect of Cultivar and Growth Regulator on In vitro Micropropagation of Potato (Solanum tuberosum L)
The present investigation was carried out aiming to develop a technique for rapid in vitro micropropagation of potato (Solanum tuberosum L) plants. Nodal explants prepared from proliferating shoots of established axenic cultures of four potato cultivars viz Diamant, Alpha, Almera and Agria were used. Explants were incubated on agar solidified (0.8% g) Murashige and Skoog's (MS) medium containing 3% sucrose and supplemented with different concentrations of thiadizuron (TDZ) and benzylaminopurine (BA) alone or in combinations with a-naphthalene acetic acid (NAA).Cultivars studied showed wide variation in their response to the plant growth regulators, best results being obtained for the cultivar Almera. Nodal explants responses to BA and TDZ were cultivar-dependent for number of shoot per explant. The highest number (5.4 shoots/explant) of shoots per explant was obtained for Almera explant cultured on MS medium supplemented with 3.0 mg/l TDZ in combination with 0.1 mg/l NAA. Regene...
Comparison of tuber and shoot formation from in vitro cultured potato explants
Plant Cell Tissue and Organ Culture, 1998
The development of axillary buds of potato (Solanum tuberosum L.) plants, cultured in vitro, was analyzed. Depending on the composition of the culture medium, the buds developed into either tubers (medium with 8% sucrose), shoots (1% sucrose), or stolons (8% sucrose and 0.5 μM gibberellin). Endogenous sugar and starch levels, and key-enzymes involved in the conversion of sucrose to starch were determined at different stages of development. Moreover, the spatial distribution of sugar levels and enzyme activities were determined within the developing structures. Glucose and fructose decreased upon tuber formation, most noticeably in the swelling parts, where also starch accumulated. The activities of sucrose synthase, fructokinase and ADP-glucose pyrophosphorylase were highest under tuber-inducing conditions, the increase being confined to the tubers, and absent in the subtending stolons. It is concluded that changes in the measured parameters, observed under tuberizing conditions, are specifically related to the formation of the tuber, and are confined to the swelling part only.
2012
A high number of micro shoots (21 +/- 2.31) of Brassica oleracea var. botrytis (cauliflower) were obtained when hypocotyl explants from 2-week-old aseptic seedlings were cultured on MS medium supplemented with 0.1 mg/L NAA and 5 mg/L BAP. Artificial or synthetic seeds were formed when the micro shoots were encapsulated in 4% (w/v) sodium alginate with 100 mM CaCl2 as complexing solution. The artificial seeds took 12 days (after 7 days storage) and 14 days (after 30 days storage) to germinate on MS basal medium. The germination percentage of artificial seeds was enhanced by the inclusion of 0.3 mg/L NAA and 3.0 mg/L BAP in the encapsulation matrix after 7 and 30 days of pre-germination storage. The time taken for germination was also faster (5 days after 7 days of storage and 11 days after 30 days of storage) when MS fortified with 0.3 mg/L NAA and 3.0mg/L BAP were used. Isolated micro shoots encapsulated in MS supplemented with 0.3 mg/L NAA and 3.0mg/L BAP gave high germination percentages (70 +/- 5.43% and 63.33 +/- 4.17%) after 7 and 30 days of pre-germination storage period, respectively. Complete plant regeneration was achieved from the germination of these artificial seeds derived from the micro shoots after 3-4 weeks in culture.