ResearchPDF Available

DKW emerges as a superior media factor in in vitro plant regeneration

Authors:
Shafkat Shamim Rahman at University of Dhaka
Shafkat Shamim Rahman
Download file PDF
Read file
Download citation

Abstract

Plant regeneration heavily depends on the basal media factor. Recent studies showed the potential role of Driver and Kuniyaki Walnut (DKW) medium as an alternative to popular MS medium. In these studies, DKW distinctly depicted superiority over MS medium in terms of multiple shoot induction and callus mediated shooting of C. ternatea and B. australis. Earlier reports also claimed, the successful regeneration studies of Pimpinella pruatjan, Theobroma cacao, Manihot esculenta Crantz, Corylus avellana, Juglans regia, Juglans nigra with a high percentage of germination and plantlet conversion in DKW medium. The disparity between DKW and MS media is mostly in their macronutrients composition, but the solidifying agent Gelrite in DKW is also assumed to be a key factor to separate them in terms of success.
Content uploaded by Shafkat Shamim Rahman
Author content
All content in this area was uploaded by Shafkat Shamim Rahman on Mar 01, 2018
Content may be subject to copyright.
DKW emerges as a superior media factor in
in vitro
plant regeneration
Shafkat Shamim Rahman*
Rahman SS. DKW emerges as a superior media factor in in vitro plant
regeneration. J Agri. 2018;1(1):3-4.
Plant regeneration heavily depends on the basal media factor. Recent
studies showed the potential role of Driver and Kuniyaki Walnut (DKW)
medium as an alternative to popular MS medium. In these studies, DKW
distinctly depicted superiority over MS medium in terms of multiple shoot
induction and callus mediated shooting of C. ternatea and B. australis.
Earlier reports also claimed, the successful regeneration studies of
Pimpinella pruatjan, Theobroma cacao, Manihot esculenta Crantz,
Corylus avellana, Juglans regia, Juglans nigra with a high percentage of
germination and plantlet conversion in DKW medium. The disparity
between DKW and MS media is mostly in their macronutrients
composition, but the solidifying agent Gelrite in DKW is also assumed to
be a key factor to separate them in terms of success.
Key Words: Plant regeneration, DKW, Gelrite
INTRODUCTION
Plant regeneration studies have discovered many biotic and abiotic
influencing factors [1]. The most important of them may be the basal
media factor. MS media [2] is the most popular due to its efficiency [3],
but a recent study also divulged the potential of another medium Driver
and Kuniyaki Walnut (DKW) [4] once again. In this study, DKW
delineated superiority over MS medium in terms of multiple shoot
regeneration, and callus mediated shooting of C. ternatea by highest
73.33% regeneration on both occasions, against 66.67% response [5]. The
study was supported by the higher multiple shoot induction from apical
buds (highest 100% DKW+ five different conc. of BA and iron as
FeEDDHA; 88% MS+8.8 μM BA) and nodal segments (highest 100%
DKW+ four different conc. of BA and iron as FeEDDHA; 94% MS+4.4
μM BA) of B. australis in DKW compared to MS [6]. Additional
micronutrients or extra vitamins in DKW may be the decisive factor
according to the author.
Before that, Deng and Cornu reported 45% walnut somatic embryo
germination to plantlet in an earlier study [7]. Somatic embryogenic
regeneration study of Pimpinella pruatjan also showed an increased
number regenerated explants [8]. Contemporary cacao (Theobroma cacao
L.) regeneration also showed high percentage successful somatic embryo
formation (5.6-66.7%), germination (50%) and plantlet conversion (65%)
in DKW medium supplemented with kinetin [9]. Nigerian cassava
(Manihot esculenta Crantz) regeneration was moderately successful with
mixed results varied from primarily 10-80% and 49-78% in further
screening [10]. Nutrient modification in DKW resulted in better
micropropagation growth for hazelnut (Corylus avellana L.) [11].
Introduction of Ni in the medium altered other nutritional requirements.
As a high salt basal medium DKW differs from others in the components
of chemical macronutrients or minerals [12]. And found superior [13] than
other media for shoot multiplication of Persian walnut (Juglans regia L.)
[14] and Juglans nigra L. (black walnut) [15].
It can be postulated the solidifying agent Gelrite in DKW play a vital role
to obtain supremacy over other basal media [16,17]. The Sphingomonas
elodea produced polysaccharide ensure the decreased use of agar [18,19]
and showed enhanced cytokinin sensitivity in bud formation of
Physcomitrella patens (Moss) [20]. But, the hypothesis on better
performance of DKW required being strengthened by supportive results of
more regeneration studies on different species.
ACKNOWLEDGEMENTS
Sincere appreciation goes to Mohammad Khairul Islam Sarkar and Chapol
Kumar Roy for their motivation and support.
CONFLICT OF INTEREST
The author declares that there is no conflict of interest exists.
ETHICS OF HUMAN AND ANIMAL EXPERIMENTS
Not applicable.
FUNDING
Not applicable.
REFERENCES
1. Shahzad A, Sharma S, Parveen S, et al. Historical Perspective and
Basic Principles of Plant Tissue Culture. Plant Biotechnology.
2017:1-36.
2. Murashige T, Skoog F. A revised medium for rapid growth and
bioassays with tobacco tissue cultures. Physiol Plant. 1962;15:473–
97.
3. Sarkar MKI, Jahan MAA, Roy CK, et al. In vitro Shoot Micro
Propagation of Medicinal Applications and Ornamental Value of
Cestrum nocturnum L. Indian J Sci Technol. 2016;9(15).
4. Driver JA, Kuniyuki AH. In vitro propagation of Paradox walnut
Juglans hindsii × Juglans regia rootstock. HortScience.
1984;19:507-9.
5. Roy CK, Mohiuddin TM, Rahman SS. Establishment of In vitro
micropropagation and callus formation protocols for large scale
production of Clitoria ternatea L. Zeitschrift für Arznei- und
Gewürzpflanzen J Med Spice Plants. 2017;22(3):136-44.
6. Padmanabhan P, Shukla MR, Sullivan JA, et al. Iron supplementation
promotes in vitro shoot induction and multiplication of Baptisia
australis. Plant Cell Tiss Organ Cult. 2017;129:145–52.
7. Deng MD, Cornu D. Maturation and germination of walnut somatic
embryos. Plant Cell, Tiss Organ Cult. 1992;28:195-202.
8. Roostika I, Purnamaningsiha R, Darwatib I, et al. Regeneration of
Pimpinella pruatjan Through Somatic Embryogenesis. Indo J Agri
Sci 2007;8(2):60-66.
Leer to editor
United Surgical (BD) Ltd. Plot# 659-661, Islampur, Kadda, Gazipur-1702, Bangladesh
*Correspondence: Shafkat Shamim Rahman, United Surgical (BD) Ltd. Plot# 659-661, Islampur, Kadda, Gazipur-1702, Bangladesh, Tel:
+8801912731521, E-mail: shafkatshamimrahman@gmail.com
Received: January 4, 2018, Accepted: January 8, 2018, Published: January 24, 2018
This open-access article is distributed under the terms of the Creative Commons Attribution Non-Commercial License (CC BY-NC) (http://
creativecommons.org/licenses/by-nc/4.0/), which permits reuse, distribution and reproduction of the article, provided that the original work is
properly cited and the reuse is restricted to noncommercial purposes. For commercial reuse, contact reprints@pulsus.com
J Agri Vol.1 No.1 2018 3
9. Ajijah N, Hartati RS, Rubiyo, et al. Effective cacao somatic embryo
regeneration on kinetin supplemented DKW medium and somaclonal
variation assessment using SSRs markers. AGRIVITA. 2016;38(1):
80-92.
10. Nkaa FA, Ene-Obong EE, Taylor NJ, et al. Evaluation of Ten Elite
Nigerian Cassava (Manihot esculenta Crantz) Cultivars for Somatic
Embryogenesis and Regeneration Potentials. J Adv Biol Biotechnol
2015;2(2):133-144.
11. Hand C, Reed BM. Minor nutrients are critical for the improved
growth of Corylus avellana shoot cultures. Plant Cell, Tiss Organ
Cult. 2014;119(2), 427-39.
12. Avilés F, Ríos D, González R, et al. Effect of Culture Medium in
Callogenesis from Adult Walnut Leaves (Juglans regia L.). Chilean J
Agri Research. 2009;69(3):460-467.
13. Payghamzadeh K, Kazemitabar SK. Comparison effects of MT novel
medium with modified DKW basal medium on walnut
micropropagation. Proceeding book of the 1st national conference of
student biology and modern world. 2008;204.
14. Saadata YA, Hennerty MJ. Factors affecting the shoot multiplication
of Persian walnut (Juglans regia L.). Sci Horticult. 2002;95:251-60.
15. Heile-Sudholt C, Huetteman CA, Preece JE, et al. In vitro embryonic
axis and seedling shoot tip culture of Juglans nigra L. Plant Cell Tiss
Organ Cult. 1986;6:189-97.
16. Payghamzadeh K, Kazemitabar SK. In vitro propagation of walnut -
A review. African J Biotechnol. 2011;10(3):290-311.
17. Huang LC, Chi DL. Pivotal roles of picloram and gelrite in banana
callus culture. Environ Exp Bot. 1988;28(3):249-58.
18. Kang KS, Colegrove GT, Veeder GT. Deacetylated polysaccharide
S-60. U.S. patent US4326052 A, 1982.
19. Kang KS, Veeder GT. Polysaccharide S-60 and bacterial
fermentation process for its preparation. U.S. patent US4326053 A,
1982.
20. Hadeler B, Scholz S, Reski R. Gelrite and Agar Differently Influence
Cytokinin-Sensitivity of a Moss. J Plant Physiol. 1995;146(3):
369-71.
Shafkat Shamim Rahman.
4J Agri Vol.1 No.1 2018
... Rahman [22] studied the DKW medium and MS medium in which he found that DKW clearly showed dominating performance over MS medium in various tissue culture stages, such as callus mediated shooting and multiple shoot induction in Clitoriaternatea and Baptisia australis. Yegizbayevaet al. [23] also reported the best results on corrected DKW formulations for proliferation, rooting or ex vitro survival of walnut genotypes. ...
... These exudates, including juglone, in the medium interrupt cell growth, inhibit growth of the explant and also adversely affect their survival [26,54,55]. The superiority of the DKW medium over the MS medium could be attributed to its distinct composition of macronutrients, and also the Gelrite (gelling agent) in DKW is thought to be one of the key factors [22]. Compared to forced shoot tips, unforced shoot tips resulted in maximum percentage of explant and media browning and a lower survival percentage. ...
In Vitro Prevention of Browning in Persian Walnut (Juglans regia L.) cv. Sulaiman
Article
Full-text available
  • Aug 2022
The present investigation was undertaken to standardize the media and the anti-browning regime in order to minimize the phenolic browning of an in vitro culture of Persian walnut cv. Sulaiman. The experiments involved two types of explants, forced and unforced shoot tips, two types of media, Driver and Kuniyuki Walnut (DKW) medium andMurashige and Skooģs (MS) medium, and three types of anti-browning agents, namely, Polyvinylpyrrolidone, ascorbic acid and activated charcoal at 150, 350 and 550 mg/L each. The investigation was replicated thrice under a completely randomized design. Forced shoot tips of cv. Sulaiman on DKW medium showed the best performance in terms of least browning (13.6 ± 10.5%) and highest survival percentage of explants (74.5 ± 2.4%) when treated with ascorbic acid at 550 mg/L. However, unforced shoot tips in MS medium did not perform well and manifested maximum browning (52.9 ± 5.2%). Based on the results, we conclude that incorporation of ascorbic acid in the DKW medium significantly reduced the media and explant browning, thus, it could set the basis of successful in vitro-propagation of walnuts.
View
... Furthermore, there are no reports on using other media, such as BLT and DKW, on turmeric growth and development under in vitro cultures. In our first two experiments, the DKW was the best medium for 'Hawaiian Red' turmeric with the highest number of complete plantlets (with roots), mainly due to differences in macronutrients such as lower nitrogen and higher sulfate concentrations than MS (Phillips and Garda, 2019;Rahman, 2018). This is evident by the high salt stress symptoms, such as yellowish leaves (pictures not shown) in the first week of transfer to this medium. ...
... The results also suggest that the DKW medium was the best performing medium. Rahman (2018) reported the differences between DKW and MS in their macronutrients composition, since DKW has lower nitrogen and higher sulfate content in its formula (Phillips and Garda, 2019). DKW has the potential to produce millions of turmeric plants within a year, and was the best medium for 'Hawaiian Red' turmeric. ...
Optimizing Turmeric Tissue Culture, Testing Different Media and a Plant Growth Regulator Matrix
Article
Full-text available
  • Aug 2021
Rapid multiplication of turmeric ( Curcuma longa ) by micropropagation is needed to produce a continuous source of uniformly sized, high-quality, and disease-free plantlets. Three in vitro experiments were conducted to optimize the medium by evaluating nine media and a full factorial combination (matrix) of two plant growth regulators for direct organogenesis of ‘Hawaiian Red’ turmeric. Two experiments evaluated the media, and the third studied the plant growth regulator matrix. As a result, Driver and Kuniyuki walnut (DKW), Murashige and Skoog (MS), and broadleaf tree basal (BLT) media performed better than woody plant media [Lloyd & McCown woody plant basal medium (L&M), and McCown’s woody plant basal salt mixture (McCown)] for shoot and root formation. The multiplication rate was 18 plants per explant in DKW with 1 mg⋅L ⁻¹ 6-benzylaminopurine (BAP) and 0.1 mg⋅L ⁻¹ 1-naphthaleneacetic acid (NAA). After transferring the plants to an ex vitro environment, the survival rate was 97%, and 30% higher than previously reported. DKW produced the highest number of plantlets (with shoots and roots), and BLT produced fewer plants with higher biomass. In the MS media, higher BAP to NAA ratio (2.5 to 0.1 mg⋅L ⁻¹ ) produced the most significant number of shoots; however, the lowest concentration of BAP and NAA (0.1 mg⋅L ⁻¹ of both) produced the highest number of rooted plantlets. There are two recommendations for tissue culture of ‘Hawaiian Red’ turmeric. To produce the highest number of plantlets, one should use the higher BAP to NAA ratio (2.5 mg⋅L ⁻¹ BAP and 0.1 mg⋅L ⁻¹ NAA) for shoot proliferation and then transfer the explants to the root initiation media. However, to reduce the number of subcultures, the explants can be grown in the lowest concentration of both BAP and NAA (0.1 mg⋅L ⁻¹ ) to induce both shoot and root. Although, the number of plantlets (with roots and shoots) will decrease in this method, there is no need for subsequent subcultures and changing of the plant growth regulator combinations.
View
... Apart from Calcium, other macroelements include magnesium, potassium, sodium, chloride, sulfate, and micronutrients: Mn, B, Cu, Ni, and Zn. The highest micronutrient content is manganese (198µM), and the lowest micronutrient content is nickel (0.02µM) [15]. ...
Growth response of four accessions of Moringa oleifera Linn shoots cultured on various basic media
Article
Full-text available
  • May 2021
Moringa ( Moringa oleifera Linn) has a highly considerable nutritional value. The conventional propagation of Moringa has limitation. Propagation through tissue culture offers uniform transplants, and free of pests and diseases. Selection of basic medium is a critical point to find out the best medium compositions for micropropagation. The research was aimed to investigate growth response of M. oleifera shoots cultured on several basic media. The experimental design was factorial complete random design with two factors i.e. types of basic medium (MS, WPM, DKW, NN, and B5) in combination with accessions of M. oleifera : Bogor, Pekalongan, Bima and Blora. Each treatment consisted of 12 replicates. The growth variables observed were shoot height, number of shoots, number of petioles, number of roots, fresh weight, dry weight of plantlets, and plantlet performance. The growth observation was carried out every week. The results showed that type of basic medium significantly affected shoot height, number of petioles, fresh weight, dry weight, and number of shoots, but, it did not significantly affect the number of roots. The accessions significantly affected shoot height, number of petioles, number of roots, fresh weight, and dry weight, but, did not significantly affect number of shoots. The ANOVA showed an interaction between basic medium in combination with accessions on all growth variables observed. The highest shoot height (13,97 cm), number of petioles (12,33), and number of roots (8.33) were achieved on DKW basic medium on Bima and Pekalongan accessions, while the highest number of shoots (6.33) was found in NN medium.
View
... Дл я культивування евкомії в'язолистої використовували живильні середовища Мурасіге-Скуга (MS) і Драйвера (DKW) (Rahman, 2018) з додаванням до їх складу цитокінінів і ауксинів: 6-БАП (6-бензиламінопурин), НОК (α-нафтилоцтова кислота) та ІОК (β-індоліл-3-оцтова Лісове господарство кислота) як окремо, так і комбінуючи їх. Додатково до середовища додавали сахарозу (0,3 %), мезоінозит (100 мг/л) та агар (0,7 %), рН середовища -5,7-5,8. ...
View
INFLUENCE OF NUTRIENT MEDIUM COMPOSITION ON REGENERATION OF SOME CLEMATIS L. CULTIVARS IN VITRO
Article
  • Dec 2023
The aim of the work was to optimize cultivation techniques at the stage of actual micropropagation of Clematis L varieties. N.V. The studies were carried out in 2021–2022 in the laboratory of plant biotechnology of N.V. Tsitsin Main Botanical Garden of the Russian Academy of Sciences (GBS RAS). Varieties of various garden groups of clematis were chosen as objects: Princess Diana, Madame Julia Correvon, Yubilejnyj 70, Multi Blue. In the course of the study, the influence of genetic characteristics and the composition of the nutrient medium on the regeneration potential of Clematis L. was shown. It was established that on the nutrient medium DKW (Driver and Kuniyuki, 1984) with the addition of 0.1 mg/l BA (6-benzylaminopurine), Clematis L. regenerants were characterized by active growth of microshoots and the highest height. It was 5.0 ± 0.2 cm for Princess Diana, 6.3 ± 0.1 cm for Madame Julia Correvon, 2.1 ± 0.1 cm for Yubilejnyj 70, and 2.8 ± 0.1 cm for Multi Blue. The maximum multiplication factor was noted for the Princess Diana variety (3.7 ± 0.1), the minimum for the Multi Blue variety (2.4 ± 0.1). When studying the effect of growth regulators and their concentrations during cultivation on DKW medium, it was shown that 2-iP (2-isopentyladenine) at a concentration of 1.5 and 2.0 mg/l was more effective than BA at the same concentrations. At the same time, the highest height (8.6 ± 0.2 cm) and multiplication factor (6.7 ± 0.2) were observed in the Madame Julia Correvon variety on a nutrient medium with the addition of 1.5 mg/l BA. The multiplication factor of variety Yubilejnyj 70 reached its maximum at 2.0 mg/l BA (4.2 ± 0.1), variety Multi Blue – at 1.5 and 2.0 mg/l 2-iP (4.0 ± 0 .1 and 4.3 ± 0.1, respectively).
View
In vitro propagation of walnut varieties from Uttarakhand
Article
Full-text available
  • Nov 2024
The Persian walnut, or English walnut, is one of the most prized walnuts, popular for its timber, medicinal properties, oil, and nuts. Due to its high demand across the globe, walnut cultivation has exponentially grown over the years. Despite having the appropriate climate, the state of Uttarakhand still does not cultivate walnut on a large scale. The alternative to growing walnuts with better quality and high production is through in vitro propagation. The present study was conducted to propagate walnut varieties growing in the Uttarakhand region through direct and indirect organogenesis. The DKW medium proved to be the most effective for nodal explant culture, while the MS medium yielded superior results for callus induction. We also optimized the media by varying the liquid/semisolid medium and plant growth regulator combinations. The genotype from higher altitudes of Uttarakhand was found to respond better to the in vitro treatments. The findings can prove useful in propagating good-quality walnut varieties in Uttarakhand on a large scale.
View
EVALUATION OF IN VITRO PROTOCOLS FOR EFFECTIVE REGENERATION OF WEST AFRICAN Theobroma cocoa (L)
Article
Full-text available
  • Nov 2024
Cacao is a perennial tree crop from the Malvaceae family. It is made up of twenty-one species, with Theobroma cacao as the commonest and one of the most economically important crops globally. West Africa is the largest region of producers. Lack of availability of planting materials all year round has been a major factor besetting cocoa production in West Africa and has led to a drastic reduction in production in the last decade. Establishing an efficient in vitro protocol for germinating zygotic embryos of local West African cocoa varieties offers a solution to expedite large-scale planting material production to enhance cocoa sustainable production and material availability. An effective in vitro protocol for germinating zygotic embryos of local West African cocoa varieties will help produce large planting materials within the shortest possible time and promote sustainable supply, which could boost cocoa production within the region and increase the availability of planting materials. Four West African local varieties of Theobroma cacao were cultured in three explant types on both Driver and Kuniyaki Walnut (DKW) basal salts and Murashige and Skoog (MS) culture media without growth regulators for twenty days. The three explant types included seeds with mucilage (SWM), seeds without mucilage (SWtM) and embryo axis (EA). The cultured explants were kept in a growth room of light intensity with a temperature of 25 ℃ ±1, relative humidity of 85% and the three explant types responded differently. Germination responses significantly varied among the explant types. SWtM sprouted earlier with more than 160 plantlets before day 5, followed by EA with 150 plantlets before day 5, while SWM showed the lowest mean germination percentage of 8% compared to SWtM and EA with 9% Early sprouting was observed in the DKW medium with almost 250 (34.72%) plantlets development before day five compared to the MS medium with less than 150 (20.83%) plantlets development, but the MS medium produces the optimal growth performance with the best mean germination time of 0.08 per day, final germination percentage of 71.39%, and development of the growth parameters leading to the development of plantlets, including 2.60 for number of leaves. This study shows that the media for in vitro culture and the source of explants significantly influence seed germination and produce dissimilar effects on the germination of zygotic embryos of Theobroma cacao varieties. The study recommends using SWtM on DKW medium for early sprouting of seeds of Theobroma cacao followed by a transfer into MS medium for seedling development into plants.
View
Iron supplementation promotes in vitro shoot induction and multiplication of Baptisia australis
Article
Full-text available
  • Apr 2017
  • PLANT CELL TISS ORG
A simple and efficient system was developed for the rapid in vitro multiplication of Baptisia australis using nodal explants derived from axenic cultures. Among the four cytokinins [benzylaminopurine (BA), kinetin, N6-(2-isopentenyl)adenine (2-iP) and zeatin] tested for axillary bud induction, BA at 4.4 µM proved to be the most effective treatment for induction of multiple shoots from nodal explants after six weeks. Supplementation of Murashige and Skoog (MS) and Driver and Kuniyuki walnut (DKW) media with a chelated iron source, ethylenediamine Bis (2-hydroxyphenylacetic acid) (FeEDDHA) and BA had a favorable effect on axillary bud proliferation from explants (apical buds and nodal segments) further improving the regeneration response as well as number of shoots induced. DKW media proved to be superior to MS for multiple shoot induction of B. australis. A combination of 4.4 µM BA and 23 µM of FeEDDHA in DKW produced a positive response resulting in 100% of nodal segments responding and regenerating 7.2 shoots per explant. The highest rooting percentage was observed on MS media containing 1 µM IBA. In vitro-rooted plants were successfully acclimated in a greenhouse with a survival rate of 97%. The micropropagation method described in this study has a potential for commercial multiplication of this horticultural plant.
View
In Vitro Propagation of Paradox Walnut Rootstock
Article
Full-text available
  • Aug 1984
Apical and lateral meristems of Paradox walnut ( Juglans hindsii x J. regia ) were used to investigate the possibility of accomplishing in vitro propagation of Paradox rootstock. A walnut specific medium, DKW, has been defined supporting optimum multiple shoot development under 4.5 μm benzyladenine (BA) and 5 nm indolebutyric acid (IBA) treatment. The method of analysis to determine the optimum level of constituents in tissue culture medium is discussed. Also reported is the in vivo rooting of tissue culture derived shoots.
View
Effective cacao somatic embryo regeneration on kinetin supplemented DKW medium and somaclonal variation assessment using SSRs markers
Article
Full-text available
  • Feb 2016
This study aimed to develop the cacao (Theobroma cacao L.) in vitro regeneration system through somatic embryogenesis on kinetin supplemented DKW medium and somaclonal variation assessment using SSR markers. Callus were initiated from basal petal and staminoid explants cultured on callus induction (CI) medium contained DKW basalt salts and kinetin:2,4-D ratios of 1:15.5, 1:7.8 or 1:3.9 and then transferred onto secondary callus growth (SCG) medium contained WPM basalt salts and kinetin:2,4-D ratios of 1:7.8 or 1:3.9. The calli were then subsequently transferred onto embryo development medium contained DKW basal salts with or without the addition of amino acids, adenine or activated charcoal for the formation of somatic embryos. Nine cacao genotypes were tested for their ability to develop somatic embryos. Results of this study indicated DKW medium supplemented with Kinetin in combination with 2,4-D effectively induced cacao somatic embryogenesis. The highest somatic embryos formation was abtained from kinetin:2,4-D ratio of 1:3.9 and 1:7.8 in CI and SCG medium respectively. Cacao genotype responses were highly explant type dependent. The developed method resulted in a high percentage of somatic embryo formation (5.6-66.7%), germination (50%) and plantlet conversion (65%) and a medium percentage of somaclonal variations based on SSRs marker analysis.
View
Effect of culture medium in callogenesis from adult walnut leaves (Juglans regia L.)
Article
Full-text available
  • Jul 2009
  • CHIL J AGR RES
To define preliminary aspects of walnut (Juglans regia L.) callogenesis in indirect propagation systems, this work analyzed callogenic induction in adult leaves of walnut in broadleaf trees (BTM), Murashige and Skoog (MS) Driver Kuniyuki Walnut (DKW) and Woody Plant (WPM) media. Leaf segments of 1 cm 2, previously sterilized, were placed in each culture medium, supplemented with 21.4 μM naphtalene acetic acid (NAA) and 8.8 μM 6-benzylaminopurine (BAP). A completely random design was used; the experimental unit was a Petri dish with six callus explants. Each treatment was composed of seven Petri dishes. Callogenesis (%), nodular calli (%) and histology of calli explants were evaluated at 4 weeks by staining 10 μm slices with safranine-fastgreen. There were no significant differences (α = 0.05) in callogenesis among evaluated media; but nodular calli percentages were significantly higher in BTM (75%) and WPM (63%). Only in calli obtained in the BTM medium did the histology show the presence of meristematic zones, but without external expression. These results indicate that the medium influences the morphogenic characteristics of resultant callus. Calli produced in BTM presented better morphogenic potential, with meristems at primary stages of organization.
View
In vitro propagation of walnut -A review
Article
Full-text available
  • Jan 2011
  • AFR J BIOTECHNOL
In vitro propagation of walnut has played a very important role in rapid multiplication of cultivars with desirable traits and production of healthy and disease-free plants. During the last several years, different approaches have been made for in vitro propagation of walnut. Micropropagation using apicale bud, nodale segement, leaves, petioles, cotyledons, embryos and understanding the specific requirement at different stages has been comprehensively covered in literature. New challenges for refinements of protocols for high rate of shoot multiplication and development of cost effective methods has gained importance in the recent past. Importance of liquid and solid static culture for callus induction, embryogenesis, shoot proliferation and root induction for walnut is also discussed in the present review. Further, the development of protocols for in vitro propagation, culture nodal segment from seedling, somatic embryogenesis and plant regeneration which is considered the most important step for successful implementation of various biotechnological technique used for plant improvement programmes has been adequately addressed in literature. In walnut, there are several reports which indicate rapid regeneration and multiplication through organogenesis or somatic embryogenesis. On the whole, the present review gives a consolidated account of in vitro propagation in walnut.
View
Minor nutrients are critical for the improved growth of Corylus avellana shoot cultures
Article
Full-text available
  • Nov 2014
Many hazelnut (Corylus avellana L.) cultivars fail to thrive in vitro on standard growth medium and the reasons for poor growth are not well understood. Our initial study of five C. avellana cultivars showed that changes in the mineral nutrients of Driver and Kuniyuki walnut (DKW) medium, including doubling the minor nutrients, produced improved growth and shoot quality. The objectives of this study were to determine the effects of the individual minor mineral nutrients from DKW medium and if added nickel was required for optimal growth. Five factors were tested at 0.5 × to 4× DKW medium concentrations, [H3BO3, CuSO4·5H2O, MnSO4·H2O, Na2MoO4·2H2O and Zn(NO3)2·6H20], in a response surface design with 39 treatment combinations. Ni was not present in the DKW medium formulation so NiSO4·6H2O was varied from 0 to 6 µM. There were many significant interactions among the minor nutrients. Higher concentrations (4×) of B, Mo, and Zn increased overall shoot quality, length, and multiplication. Increased Mo improved some responses for each cultivar, and it interacted significantly with Cu and Zn. The addition of Ni greatly improved the shoot quality and length of ‘Sacajawea.’ Ni interactions were significant for the other cultivars as well, and altered the requirements for the other minor nutrients, but did not necessarily improve the overall shoot response. Improved growth and shoot quality for most cultivars required increased amounts of B, Mo, and Zn and less Mn and Cu. ‘Sacajawea’ required increased B, Cu, Zn, and Ni. All of the cultivars required minor nutrient formulations that differed greatly from DKW medium or other published minor nutrient formulations.
View
Establishment of In vitro micropropagation and callus formation protocols for large scale production of Clitoria ternatea L.
Article
  • Nov 2017
Background and Objective: Clitoria ternatea is a local, valuable medicinal plant containing active major phytochemical compounds. Aim of this study was to establish an in vitro micropropagation protocol to produce C. ternatea at large scale. Materials and Methods: Cotyledonary nodal segments and cotyledons from aseptic seedling were used as explants for regeneration of multiple shoot plus callus induction and root formation. DKW, MS and half strength MS media supplemented with various concentrations of 6-benzyl amino purine (BAP), indole-3 acetic Acid (IAA), napthalene acetic acid (NAA) and indole-3 butyric Acid (IBA), were used as single or in combinations. Results: The highest frequency (73.33%) of shoot regeneration was obtained in DKW medium supplemented with BAP (1.0 mg/L). Maximum (80%) callus formation was observed on MS medium supplemented with BAP and NAA. Best callus mediated shooting (90%) was found from DKW medium supplemented with BAP and NAA and mean number of multiple shoot was 5.22±0.46. MS medium supplemented with IAA gave the highest frequency (90%) of root formation. Conclusion: It can be concluded that an alternative medium has equal potential to be used in future in vitro regeneration of plants, which is corroborated by the successful establishment of regeneration protocol for large scale production of Clitoria ternatea L.
View
REGENERATION OF Pimpinella pruatjan THROUGH SOMATIC EMBRYOGENESIS
Article
  • Oct 2016
Pruatjan (Pimpinella pruatjan Molk.) is an Indonesian endangered plant which has various medicinal properties such as aphrodisiac, diuretic, and tonic. The plant is commonly harvested from its natural habitat, therefore it becomes endangered. Regeneration of pruatjan through organogenesis has been studied, but its shoot multiplication was very low (5 shoots per explant). The study aimed to investigate the best regeneration technique of pruatjan through somatic embryogenesis. This research was conducted at the tissue culture laboratory, Indonesian Center for Agricultural Biotechnology and Genetic Resources Research and Development in 2004-2005. Callus formation of pruatjan was induced from the petioles and leaves in Driver and Kuniyaki’s (DKW) based medium containing 2,4-D combined with picloram at the level of 0.5, 1.0, 1.5, and 1.5 ppm. Embryogenic calli were then transferred into embryo development medium in two ways. First, they were directly transferred into media containing IBA/NAA at the level of 0.5, 1.0, and 1.5 ppm. Second, they were indirectly transferred into media containing 2.0 ppm 2,4-D and 0.3% casein hydrolysate prior to the IBA/ NAA media. Parameters evaluated were fresh weight, dry weight, time initiation of embryogenic callus formation, and total number of embryos. The result showed that calli of pruatjan were successfully induced from the petioles and leaves. The best calli were induced from the leaves in the DKW medium containing 2.0 ppm 2,4-D and 0.5 ppm picloram. Embryo development of the calli was best if they were first grown in the media containing 2.0 ppm 2,4-D and 0.3% casein hydrolysate then transferred to the IBA/NAA media. The total number of somatic embryos was counted up to 103 on the medium containing 1.5 ppm IBA. This study indicated that pruatjan somatic embryogenesis regeneration required three different media, i.e. for callus induction, development and maturation, and for germination.
View
View
View