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1.
Theoretically, complete rejuvenation of mature trees should occur through somatic embryogenesis, however, this has not been
extensively studied. The main objective of the present study was to increase the efficiency of in vitro clonal propagation
for mature Quercus robur (100–300 years old), by induction of somatic embryogenesis as rejuvenation step prior to establishment of shoot culture through
micropropagation of somatic embryo-derived plantlets. Shoot culture lines of “mature” origin were established from epicormic
shoots of two centenarian oak genotypes (Sainza and CR-0) and maintained by axillary shoot proliferation. Embryogenic lines
were also initiated from epicormic leaf explants of the same genotypes and maintained by secondary somatic embryogenesis.
Although the frequency of somatic embryo conversion into plantlets was low in pedunculate oak, shoot culture lines could be
established and maintained by axillary branching from several germinated somatic embryos. For each genotype and shoot culture
line of the two origins (mature tree and somatic plantlets), shoot multiplication rate and elongation as well as rooting ability
parameters were compared. Compared with “mature-origin” shoot cultures and after more than one year propagation in vitro,
shoot lines established from somatic plantlets produced a significantly higher proportion of elongated, rootable shoots (from
26.0–31.6 to 36.8–40.5%) with increased rooting ability (from 3.3–45.6% to 23.2–89.8%). In the case of 300-year-old Sainza
genotype such a high organogenic capacity was similar to shoot cultures initiated from basal sprouts. Basal sprouts are considered
as “mature” material that retains juvenile characteristics compared with epicormic shoots forced from crown branches. Somatic
embryogenesis only slightly improved plant regeneration of shoot cultures from basal sprouts, thus validating their use as
“juvenile control”. The present results provide evidence that some rejuvenation occurred during the process of somatic embryogenesis
and resulted in improved shoot growth and rooting of somatic embryo-derived culture compared with “mature” shoot culture.
The results reported in this study might be useful in embryogenic systems with low plant conversion rates. The proposed experimental
model might also be useful in finding molecular markers of plant ontogeny. 相似文献
2.
Haoru Tang Zhenglong Ren Gari Krczal 《In vitro cellular & developmental biology. Plant》2000,36(1):47-50
Summary Well-developed somatic embryos were selected from a repetivively somatic embryo line derived from embryonic axes of immature
zygotic embryos of English walnut ‘No. 120’ (Juglans regia L.) for germination and conversion studies. In germinating dishes, somatic embryos germinated into only shoots, only roots,
or both shoots and roots. Without any pretreatment, 28% somatic embryos germinated, while those treated with 2.5–5.0 mg 1−1 (7.2–14.4 μmol) gibberellic acid (GA3) germinated at 25–28% and those receiving a cold treatment of 2–3 mo. at 3–4°C germinated at 30–43%. However, only 4–19%
of the germinating embryos showed both shoots and roots. Treated with desiccation, either with CaCl2·6H2O or Ca(NO3)2·4H2O at 20°C in the dark for 3 d, somatic embryos germinated at 85–91%, 57–69% of which had both shoots and roots. Treatment
with 2 mo. cold storage in combination with desiccation using Ca(NO3)2·4H2O resulted in 92% of somatic embryos germinating, 70% of which showed both shoots and roots. No significant differences were
observed between solid and liquid germination media. After transferring the germinating embryos to plantlet development media,
52–63% of those with both shoots and roots developed into plantlets while 11% with only shoots or 9% with only roots converted
into plantlets. Plantlet development was improved by using lower medium salts and sucrose concentrations. The addition of
activated charcoal enhanced root development, particularly root branching. Of 131 plants transplanted, 91 plants were acclimatized
to a greenhouse. 相似文献
3.
Rubén Mallón Purificación Covelo Ana María Vieitez 《Trees - Structure and Function》2012,26(3):731-741
The effects of the culture system used for embryo proliferation were investigated with the aim of improving multiplication rates and somatic embryo quality in two embryogenic lines of Quercus robur derived from mature trees (B-17 and Sainza). Embryo proliferation medium was defined following comparison of five different semi-solid media, and the highest multiplication rates (based on the total number of embryos and number of cotyledonary-shaped embryos) were achieved with medium supplemented with 0.44 μM benzyladenine for both lines. Embryo proliferation on semi-solid medium was compared with that obtained by a temporary immersion system (TIS), in which four cycles with immersion frequencies of 1 min every 6, 8, 12 or 24 h were tested. TIS promoted a significant increase in proliferated embryo biomass, with the growth index (GI) two and four times higher than in semi-solid medium in B-17 and Sainza genotypes, respectively. An immersion cycle of 1 min every 8 or 12 h produced approximately 700 somatic embryos (B-17) and 1,500 somatic embryos (Sainza) per RITA® bioreactor, with significant differences in the latter genotype with respect to gelled medium. TIS had also a significant effect on somatic embryo synchronization as it enabled a higher production of cotyledonary embryos (90%), which represents increases of 14% (B-17) and 20% (Sainza) with respect to gelled medium. For germination of embryos proliferated in TIS two maturation systems were applied: (1) culture in semi-solid medium containing 6% sorbitol or (2) culture by TIS (without sorbitol) at a frequency of 1 min immersion every 48 h. Germination ability was higher after maturation on sorbitol medium and plantlet conversion occurred in 48% (B-17) and 13% (Sainza) embryos. TIS produced large numbers of well-developed cotyledonary embryos, hence reduced the cost and labor. 相似文献
4.
Summary Cotyledonary Quercus robur L. somatic embryos from two cell lines were encapsulated in 4% (w/v) sodium alginate. An artificial endosperm was provided
by the addition of P24 medium plus 3% (w/v) sucrose. Oak somatic embryos and oak synthetic seeds were germinated on P24 medium plus 0.1 μM indole-3-butyric acid and 0.9 μM 6-benzylaminopurine or were dehydrated prior to germination. The highest conversion rates (26%) were obtained with encapsulated
somatic embryos as well as artificial endosperm-coated somatic embryos. Encapsulation improved the regeneration into oak plantlets
in one of the two cell lines tested. The artificial endosperm had no additional beneficial effect on conversion frequency,
but increased germination rate in one cell line tested. Significant higher conversion could be attributed to slow desiccation
compared to the non-encapsulated control. Cold storage as a post-maturation treatment had no influence on the germination
ability of oak synthetic seeds. Differences in the response of the cell lines with respect to conversion frequencies and timing
of germination were observed. Fifty-six well-developed plantlets regenerated 12 wk after germination, and 29 plants were transferred
to the greenhouse, where they have been successfully established in substrate. 相似文献
5.
Xueping Shi Xigang Dai Guofeng Liu Junwei Zhang Guogui Ning Manzhu Bao 《In vitro cellular & developmental biology. Plant》2010,46(2):117-125
An efficient protocol for secondary somatic embryogenesis in camphor tree is reported. Secondary somatic embryos (SSEs), initially
obtained from the primary embryos of a nascent embryogenic culture in 2002, were proliferated and maintained for more than
4 yr via cyclic secondary somatic embryogenesis. Throughout this period, the embryo populations retained a high level of competence
for plant regeneration. SSEs were produced on the surfaces of the cotyledons and radicular ends of maternal somatic embryos
(MSEs). Histological observations of the various stages of secondary embryo development revealed four typical stages, namely,
globular, heart-shaped, torpedo, and cotyledonary. The process of secondary embryogenesis continued in a cyclic way, with
each newly formed embryo producing a subsequent generation of secondary embryos. In order to progress developmentally beyond
proliferation cycles, cotyledonary embryos from one of embryogenic lines (L14) were cultured on Murashige and Skoog (MS) medium
with 0.1–3.0 mg l−1 abscisic acid (ABA) or 0.05–1.0 mg l−1 thidiazuron (TDZ) in darkness for 2 mo to achieve maturation. Matured embryos were then transferred to MS-based germination
medium containing either 0.1 mg l−1 TDZ, 0.2 mg l−1 indole-3-butyric acid (IBA), and 0.5 mg l−1 6-benzylaminopurine (BA) or 0.1 mg l−1 TDZ and 0.2 mg l−1 IBA and were cultured in light for germination. Over 50% of embryos matured in the presence of 0.5 mg l−1 ABA were able to germinate with shoots and poor root system. Frequencies of embryos germinating normal shoots among different
genotypes did not change significantly. A total of 93% of the shoots from the germinated embryos converted to plantlets on
half strength MS medium with 0.5 mg l−1 IBA by 3 wk. Plantlets acclimatized successfully to ex vitro conditions and developed as field-grown plants with normal appearance. 相似文献
6.
S. Paul A. Dam A. Bhattacharyya T. K. Bandyopadhyay 《Plant Cell, Tissue and Organ Culture》2011,105(2):271-283
A reproducible protocol for direct and indirect somatic embryogenesis was established in a small aromatic tree, Murraya koenigii. Embryogenic callus was obtained from 90% zygotic embryonic axis (ZE) and 70% cotyledon (COT) explants in Murashige and Skoog
(MS) basal medium supplemented with 8.88 μM 6-benzyladenine (BA) and 2.675 μM α-naphthaleneacetic acid (NAA). Globular somatic
embryos were induced and further matured from such embryogenic callus by subsequent culture on the same basal media containing
thidiazuron (TDZ) (2.27–9.08 μM). The highest frequency of somatic embryos (14.58 ± 0.42) was recovered from ZE-derived callus
after 6 weeks. The age and type of explant and concentration of TDZ played an important role in the development of somatic
embryos. Explants excised from 60-day-old seed differentiated from 96.67% of ZE explants and 86.67% from COT explants when
cultured on MS basal medium supplemented with 4.54 and 9.08 μM TDZ, respectively, after 4 weeks. The best result obtained
for the average frequency of somatic embryos (11.28 ± 0.32) was from ZE explants, which was significantly higher than COT
explants (7.34 ± 0.97). Most of the somatic embryos (above 95%), irrespective of their origin, germinated after 4 weeks in
1/2 MS basal media containing 2.32 μM kinetin (KN) and 1.07 μM NAA. Well-rooted plantlets were successfully acclimatized.
Histological analysis and scanning electron micrographs confirmed the initiation, development, and germination of somatic
embryos from both explants. 相似文献
7.
The production of ethylene and the endogenous content of polyamines (PAs) have been recorded during the early development,
maturation and germination of holm oak (Quercus ilex L.) somatic embryos. Ethylene production was high in embryogenic callus, immature somatic embryos and in explants showing
secondary embryogenesis, while it was lower in mature and germinating somatic embryos. A higher ethylene production was also
associated to the process of secondary embryogenesis. The exogenous application of 1-amino-1-cyclohexane carboxylic acid was
not significantly effective on the production of ethylene by holm oak somatic embryos. Total PAs were more abundant in embryogenic
callus and in both somatic and zygotic immature embryos, decreasing later on in the mature and germination phases. Immature
somatic embryos of holm oak and immature zygotic embryos contain high levels of spermidine (Spd), which decreased during maturation
and germination. Spermine (Spm) concentration was lower than that of Spd. Spm was more abundant in embryogenic callus and
immature zygotic embryos than in mature embryos. Ethylene production did not seem to interfere with PA metabolism. 相似文献
8.
Somatic embryogenesis was induced in expanding leaf explants excised from epicormic shoots forced from branch segments taken at four different times of year from a mature oak (Quercus robur L.). Branch segments 2–4 cm in diameter produced most shoots when collected in March. Somatic embryos were induced on explants derived from branches of all collection dates, although collection in November seemed to afford the best results. Germination and conversion ability of embryos of embryogenic lines derived from six oak trees depended heavily on genotype, conversion rates ranging from 0 to 70%. RAPD analyses found no evidence of genetic variation either within or between the embryogenic lines established from three of these trees, or between these lines and the trees of origin, or between somatic embryo derived plantlets and the trees of origin. The embryogenic system used in this study appears to be suitable for true-to-type clonal propagation of mature oak genotypes. 相似文献
9.
Pradeep C. Deo Robert M. Harding Mary Taylor Anand P. Tyagi Douglas K. Becker 《Plant Cell, Tissue and Organ Culture》2009,99(1):61-71
Callus was initiated in three different “esculenta” taro cultivars by culturing corm slices in the dark on half-strength MS
medium supplemented with 2.0 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D) for 20 days followed by subculture of all corm slices
to half-strength MS medium containing 1.0 mg/l thidiazuron (TDZ). Depending on the cultivar, 20–30% of corm slices produced
compact, yellow, nodular callus on media containing TDZ. Histological studies revealed the presence of typical embryogenic
cells which were small, isodiametric with dense cytoplasms. Somatic embryos formed when callus was transferred to hormone-free
medium and ~72% of the embryos germinated into plantlets on this medium. Simultaneous formation of roots and shoots during
germination, and the presence of shoot and root poles revealed by histology, confirmed that these structures were true somatic
embryos. Plants derived from somatic embryos appeared phenotypically normal following 2 months growth in a glasshouse. This
method is a significant advance on those previously reported for the esculenta cultivars of taro due to its efficiency and reproducibility. 相似文献
10.
Rubén Mallón Teresa Martínez Elena Corredoira Ana M. Vieitez 《Trees - Structure and Function》2013,27(5):1285-1296
This is the first report on the successful induction of somatic embryogenesis in swamp white oak from leaf and shoot apex explants excised from in vitro shoot cultures derived from 6- to 7-year-old trees. We demonstrated that arabinogalactan from larch wood (2–4 mg/L) promoted embryogenesis in the three genotypes evaluated by increasing the frequency of somatic embryogenesis, the embryogenic sites per explant, and by speeding the onset of embryo initiation. The explants were cultured sequentially on three culture media consisting of Murashige and Skoog (MS) salts and vitamins supplemented with 500 mg/L casein hydrolysate and different concentrations of α-naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BA). Somatic embryogenesis induction frequencies of up to 12.4, 4.5, and 0.7 % were obtained for the three genotypes. Clonal embryogenic lines were maintained by repetitive embryogenesis following culture on MS medium containing 0.44 μM BA with or without 0.27 μM NAA. Before germination, cotyledonary-stage embryos were cultured for 4 weeks in maturation medium (MS medium with half-strength macronutrients) containing 6 % sorbitol. Germination response was significantly improved by applying a 2-month cold storage as a post-maturation treatment. The mineral formulation and plant growth regulator content of the germination medium influenced the frequency of plantlet conversion with the best results achieved on Gresshoff and Doy medium with BA (0.25–0.44 μM). This procedure resulted in over 50–60 % of germinating embryos exhibiting continuous root growth and either epicotyl elongation or shoot development. 相似文献
11.
The regeneration of somatic seedlings from selected 100-year-old cork oak trees is reported. The induction of somatic embryogenesis from leaves of epicormic shoots was significantly affected by genotype, harvesting time and their interaction. Leaves from all five selected trees produced somatic embryos when the segments of branches used as sources of epicormic shoots were collected in May. Genotype, but not the level of photosynthetically active radiation, affected the proliferation of the embryogenic lines and the number of detachable embryos that could be obtained from them. Genotype also affected several steps leading to conversion of somatic embryos, from germination to complete acclimatisation of somatic seedlings. Almost 40% of the somatic embryos from all lines germinated, showing coordinated root and shoot growth. Although the mean percentage of recovery for the whole process was low, plants could be regenerated from four of the five trees tested. 相似文献
12.
Liberato Portillo Fernando Santacruz-Ruvalcaba Antonia Gutiérrez-Mora Benjamín Rodríguez-Garay 《In vitro cellular & developmental biology. Plant》2007,43(6):569-575
Somatic embryogenesis was achieved from leaves of Agave tequilana Weber cultivar azul utilizing MS medium supplemented with L2 vitamins and the addition of cytokinins: 6-benzylaminopurine
(BA), 1-phenyl-3(1,2,3-thiadiazol-5-yl)urea (TDZ), 6-(γ-γ-dimethylamino)purine (2ip) and 6-furfurylaminopurine (KIN), combined
with the auxin 2,4-dichlorophenoxyacetic acid (2,4-D). Differences among the six genotypes studied with regard to their embryogenic
response in culture were found. Embryos produced by genotype S3 under a hormone regime of high cytokinin (44.4 to 66.6 μM
BA) compared to auxin (4.5 μM 2,4-D) contained chlorophyll, whereas those produced when auxin was high compared to cytokinin
(9.0 and 13.6 μM 2,4-D and 1.3 and 4.0 μM BA, respectively) were whitish and morphologically similar to their zygotic counterparts.
Somatic embryos matured and germinated after transferring the embryogenic calli to maturation and germination medium without
growth regulators and enriched with organic nitrogen. Microscopic observations demonstrated a unicellular origin for production
of indirect somatic embryos. 相似文献
13.
S. Zdravković-Korać J. Milojević Lj. Tubić D. Ćalić-Dragosavac N. Mitić B. Vinterhalter 《Plant Cell, Tissue and Organ Culture》2010,101(2):237-244
A protocol has been developed for somatic embryogenesis and subsequent plant regeneration in Allium schoenoprasum L. Calli were induced from root sections isolated from axenic seedlings and cultivated on media containing either Murashige
and Skoog’s (MS) or Dunstan and Short’s mineral solution supplemented with 5 μM 2,4-dichlorophenoxyacetic acid (2,4-D) in
combination with 6-benzylaminopurine (BA), 6-furfurylaminopurine (Kin) or thidiazuron (TDZ) at 1, 5 or 10 μM. The highest
frequencies of callus induction were achieved on media with 5 μM 2,4-D in combination with 5 μM TDZ or 10 μM BA (78.9% and
78.4%, respectively). Calli were then transferred to 1 μM 2,4-D, where compact yellow callus turned to segmented yellowish
callus with transparent globular somatic embryos at the surface. Calli that were previously grown on media with 5 μM 2,4-D
in combination with 10 μM BA or 10 μM TDZ showed the highest frequencies of embryogenic callus formation (45% and 42%) as
well as mean number of somatic embryos per regenerating callus. The choice of mineral solution formulation did not significantly
affect callus induction or embryogenic callus formation. The embryos could complete development into whole plants on plant
growth regulator (PGR)-free medium, but inclusion of Kin (0.5, 2.5 and 5 μM) in this phase improved somatic embryo development
and multiplication. Subsequently transferred to 1/2 MS PGR-free medium, all embryos rooted and the survival rate of the plants
in a greenhouse was 96%. 相似文献
14.
Jing Li Yang Yu Da Niu Chuan Ping Yang Gui Feng Liu Cheng Hao Li 《Plant Cell, Tissue and Organ Culture》2011,106(3):391-399
Somatic embryogenesis (SE) was induced in female flower buds from mature Schisandra chinensis cultivar ‘Hongzhenzhu’. Somatic embryo structures were induced at a low frequency from unopened female flower buds and excised
unopened on Murashige and Skoog (MS) agar medium containing 4.0 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D). Friable embryogenic calli were induced from somatic embryo structures after three
to four subcultures on initiation medium. The frequencies of mature somatic embryo germination and plantlet conversion were
low, but increased in the presence of gibberellic acid (GA3). Some germinated somatic embryos could form friable embryogenic calli on medium without plant growth regulators (PGRs).
The germination and conversion frequencies of somatic embryos from embryogenic calli induced using PGR-free medium were higher
than for somatic embryos from embryogenic calli induced on medium containing 2,4-D. Most somatic embryos from 2,4-D-induced
embryogenic calli had trumpet-shaped embryos, and most somatic embryos from PGR-free medium–induced embryogenic calli had
two or three cotyledons. Histological observation indicated that two- and three-cotyledon embryos had defined shoot primordia,
but most of the trumpet-shaped embryos yielded plantlets that lacked or had poorly developed meristem tissue. Cytological
and random amplification of polymorphic DNA (RAPD) analyses indicated no evidence of genetic variation in the plantlets of
somatic embryo origin. 相似文献
15.
The induction of somatic embryogenesis from shoot apices and leaf explants of shoot cultures derived from 6- to 7-year-old
white oak (Quercus alba L.) trees is reported in this study. Embryogenic response was obtained in two out of the three genotypes evaluated with embryo
induction frequencies up to 50.7% for WOQ-1 and 3.4% for WOQ-5 genotypes. The embryogenic explants formed translucent nodular
structures and cotyledonary-stage somatic embryos, which developed from callus tissue, indicating an indirect embryogenesis
process. An efficient procedure was developed for WOQ-1 material on the basis of the most appropriate leaf developmental stage.
Growing leaves excised from two nodes below the shoot apex showed the highest embryogenic induction index. These leaves contain
cells in an undifferentiated state, as shown by the presence of precursor cells of stomata, absence of intercellular spaces
and low starch content in the mesophyll cells. Nodular structures and/or somatic embryos began to arise 7–8 weeks after culture
initiation, although most emerged after 9–12 weeks in culture. The sequence of application of media for somatic embryo induction
was optimized with a two-step procedure consisting of culturing the explants in medium supplemented with 21.48 μM NAA and
2.22 μM BA for 8 weeks and transfer of explants into plant growth regulator-free medium for another 12 weeks. Clonal embryogenic
lines were established and maintained by secondary embryogenesis. Embryo germination (30%) and plantlet conversion (16.6%)
were achieved after cold storage for 2 months. 相似文献
16.
Konstantin V. Kiselev Anna V. Turlenko Yuri N. Zhuravlev 《Plant Cell, Tissue and Organ Culture》2009,99(2):141-149
A somatic embryogenesis protocol for plant regeneration of northern red oak (Quercus rubra) was established from immature cotyledon explants. Embryogenic callus cultures were induced on Murashige and Skoog medium
(MS) containing 3% sucrose, 0.24% Phytagel™, and various concentrations of 2,4-dichlorophenoxyacetic acid (2,4-d) after 4 weeks of culture in darkness. A higher response (66%) of embryogenic callus was induced on 0.45 μM 2,4-d. Higher numbers of globular- (31), heart- (17), torpedo- (12), and cotyledon-stage (8) embryos per explant were obtained
by culturing embryogenic callus on MS with 3% sucrose, 0.24% Phytagel™, and devoid of growth regulators after 8 weeks culture
in darkness. Continuous sub-culturing of embryogenic callus on medium containing 2,4-d yielded only compact callus. Desiccation of embryos for 3 days in darkness at 25 ± 2°C followed by cold storage at 4°C in
darkness for 8 weeks favored embryo germination and development of plantlets. Cotyledon-stage embryos subjected to desiccation
and chilling treatment cultured on MS with 3% sucrose, 0.24 Phytagel™, 0.44 μM 6-benzylaminopurine (BA), and 0.29 μM gibberellic
acid germinated at a higher frequency (61%) than with 0.44 μM BA alone and control cultures. Germinated plantlets developed
a shoot and root, were acclimatized successfully, and maintained in a growth room for plantlet development. 相似文献
17.
A. Othmani C. Bayoudh N. Drira M. Marrakchi M. Trifi 《Plant Cell, Tissue and Organ Culture》2009,97(1):71-79
Plant regeneration through somatic embryogenesis from young leaf explants (5–10 mm long) adjacent to the apex of 5–6 year
old offshoots of Tunisian date palm (Phœnix dactylifera L.), cultivar Boufeggous was successfully achieved. Factors affecting embryogenic callus initiation, including plant growth
regulators and explant size, were investigated. The highest induction frequencies of embryogenic calli occurred after 6–7 months
on MS medium supplemented with 10 mg l−1 2,4-D and 0.3 mg l−1 activated charcoal. The subculture of these calli onto maintenance medium resulted in the formation of proembryos. Fine chopping
and partial desiccation (6 and 12 h) of embryogenic calli with proembryos prior to transfer to MS medium supplemented with
1 mg l−1 ABA stimulated the rapid maturation of somatic embryos. Maturated somatic embryo yield per 0.5 g FW of embryogenic callus
was 51 embryos with an average maturation time of 55 days. This was increased to 422 with finely chopped callus, and 124 and
306 embryos following 6 and 12 h desiccation treatments, respectively. The average time to maturation for these 3 treatments
was 35, 43 and 38 days, respectively. Subsequent substitution of ABA in MS medium with 1 mg l−1 NAA resulted in the germination and conversion of 81% of the somatic embryos into plantlets with normal roots and shoots.
The growth of regenerated somatic plants was also monitored in the field. 相似文献
18.
Elena Corredoira Silvia Valladares Ana M. Vieitez Antonio Ballester 《In vitro cellular & developmental biology. Plant》2008,44(4):307-315
For the mass production of chestnut trees with selected, hybrid, or genetically engineered genotypes, one potentially desirable
propagation strategy is based on somatic embryogenesis. Although methods exist for the initiation of embryogenic cultures
of Castanea sativa from immature zygotic embryos or leaf explants, the embryos produced have had low rates of conversion into plantlets. This
study explored the possible benefits for somatic embryos that have already undergone maturation and cold treatments, of (a)
partial slow or fast desiccation, and (b) of the addition of plant growth regulators or glutamine to the germination medium.
Germination response was evaluated in terms of both conversions to plantlets and through embryos developing only shoots (shoot
germination) that could be rooted following the micropropagation protocols developed for chestnut. Two or 3 wk slow desiccation
in sealed empty Petri dishes resulted in a slight reduction in water content that nevertheless increased total potential plant
recovery, shoot length, and the number of leaves per plantlet. However, best results were achieved by 2 h fast drying in a
laminar flow hood, which reduced embryo moisture content to 57–58% and enhanced the potential plant recovery and quality of
regenerated plantlets. Plant yield was also promoted by addition of 0.44 μM benzyladenine and 200–438 mg/l of glutamine to
the germination medium, and plantlet quality (as evidenced by root, shoot, and leaf growth) by the further addition of 0.49 μM
indole-3-butyric acid. 相似文献
19.
A simple and efficient system was developed for rapid somatic embryogenesis from leaf explants of Merwilla plumbea, a traditional but threatened medicinal plant in South Africa. Friable embryogenic callus (FEC) was obtained from leaf explants
on embryogenic callus induction medium containing agar-solidified Murashige and Skoog (MS) salts and vitamins, 8.3 μM picloram,
2.3 μM thidiazuron (TDZ) and 20 μM glutamine. FEC was subsequently incubated in embryogenic callus proliferation medium containing
4.5 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 4.1 μM picloram for 7 days before it was transferred to liquid somatic embryo
medium (SEML) containing MS medium supplemented with 0.4 μM picloram and 0.9 μM TDZ. In SEML supplemented with 150 mg L−1 haemoglobin, 5.4–35.6 somatic embryos per settled cell volume of 500 mg FEC were obtained. These embryos were at globular
to cotyledonary developmental stages. Embryo maturation, germination and plant formation rate was 94.4% following transfer
of SEs to half-strength MS medium supplemented with 1.4 μM gibberellic acid. Plantlets transferred into soil acclimatized
in the misthouse and established successfully in the greenhouse (100%). This is the first report on induction of Merwilla plumbea somatic embryogenesis. The protocol developed offers controlled vegetative propagation by alleviating extinction threats,
ensures germplasm conservation and provides a system for physiological, biochemical, molecular and cellular studies of embryo
development. 相似文献
20.
Jin Cui Jianjun Chen Richard J. Henny 《In vitro cellular & developmental biology. Plant》2009,45(1):34-43
Plant regeneration through direct somatic embryogenesis in Aeschynanthus radicans ‘Mona Lisa’ was achieved in this study. Globular somatic embryos were formed directly from cut edges of leaf explants and
cut ends or on the surface of stem explants 4 wk after culture on Murashige and Skoog (MS) medium supplemented with N-phenyl-N′-1, 2, 3-thiadiazol-5-ylurea (TDZ) with α-naphthalene acetic acid (NAA), TDZ with 2,4-dichlorophenoxyacetic acid (2,4-D),
or 6-benzylaminopurine (BA) or kintin (KN) with 2,4-D. MS medium containing 9.08 μM TDZ and 2.68 μM 2,4-D resulted in 71%
of stem explants producing somatic embryos. In contrast, 40% of leaf explants produced somatic embryos when induced in medium
containing 6.81 μM TDZ and 2.68 μM 2,4-D. Somatic embryos matured, and some germinated into small plants on the initial induction
medium. Up to 64% of stem explants cultured on medium supplemented with 9.08 μM TDZ + 2.68 μM 2,4-D, 36% of leaf explants
cultured on medium containing 6.81 μM TDZ and 2.68 μM 2,4-D had somatic embryo germination before or after transferring onto
MS medium containing 8.88 μM BA and 1.07 μM NAA. Shoots elongated better and roots developed well on MS medium without growth
regulators. Approximately 30–50 plantlets were regenerated from each stem or leaf explant. The regenerated plants grew vigorously
after transplanting to a soil-less substrate in a shaded greenhouse with more than a 98% survival rate. Three months after
their establishment in the shaded greenhouse, 500 plants regenerated from stem explants were morphologically evaluated, from
which five types of variants that had large, orbicular, elliptic, small, and lanceolate leaves were identified. Flow cytometry
analysis of the variants along with the parent showed that they all had one identical peak, indicating that the variant lines,
like the parent, were diploid. The mean nuclear DNA contents of the variant lines and their parent ranged from 4.90 to 4.99 pg
2C−1, which were not significantly different statistically. The results suggest that the regenerated plants have a stable ploidy
level, and the regeneration method established in this study can be used for rapid propagation of ploidy-stable Aeschynanthus radicans. 相似文献