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1.
This study established a highly effective micropropagation system to obtain good plantlet proliferation from floral organs via callus induction and bud differentiation in Guzmania ‘Hilda’ bromeliad. The best frequencies of organogenic callus formation (20% in petal and 35% in ovary explants) were obtained on media containing a combination of 1.0 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) with 1.0 mg l−1 α-naphthaleneacetic acid (NAA) and 1.5 mg l−1 2,4-D with 0.5 mg l−1 NAA, respectively. Organogenic calli were cultured on medium with 1.0 mg l−1 NAA and 0.5 mg l−1 1-phenyl-3-(1,2,3-thiadiazol-5-yl) urea (TDZ) induce the differentiation and regeneration of adventitious buds into plantlets. When the plantlets were cultured in a medium with optimum NAA concentration (0.5–1.0 mg l−1) significant improvement in regeneration and elongation was achieved within one month. This overcame the difficulty of delayed elongation in Guzmania plantlets. More than 99% of the regenerated and acclimatized plantlets developed to the flowering stage.  相似文献   

2.
To produce mass propagules of Oncidium ‘Sugar Sweet’, we tested the feasibility of producing protocorm-like bodies (PLBs) using 5 l balloon type air-lift bioreactors, and selected an optimal culture medium for shooting and rooting in vitro. The results showed that liquid bioreactor cultures were more efficient for PLBs proliferation when compared to solid- and liquid-agitated flask cultures. The maximum PLBs biomass (326.3 g per bioreactor, FW) was obtained in the immersion bioreactor culture, with the growth ratio reaching 10.9 after 50 days of culture. This was obviously higher than the ebb and flood bioreactor culture. During bioreactor culture, sucrose and electric conductivity (EC) in the culture medium were negatively correlated with PLBs growth; the highest PLBs fresh and dry biomass was obtained 40 days after culture. An inoculation density of 20 g (FW) was optimal for PLBs growth in a 3 l working volume of 5 l bioreactor. Furthermore, MS medium supplemented with 2.0 mg l−1 BA for shooting and 0.5 mg l−1 IBA for rooting was optimal during in vitro culture, the plantlets were successfully established in the potting substrate.  相似文献   

3.
Big bluestem (Andropogon gerardii Vitman) and little bluestem [Schizachyrium scoparium (Michaux) Nash.] are native to the North America and are important forage grasses and ornamental grasses. Both grasses are proposed as ideal biomass producers for cellulosic ethanol production. To apply genetic transformation, which is an important tool for incorporating desirable agronomic traits into plants to both species, however requires an efficient and reproducible regeneration protocol. We used mature caryopses from big and little bluestem as explants and tested the effect of various combinations of 2, 4-dichlorophenoxyacetic acid (2, 4-D) (1, 2, 3, 4 or 5 mg l−1) and kinetin (KT) (0, 0.1 or 0.2 mg l−1) on embryogenic callus induction with LS as the basal medium. The highest percentage of embryogenic calli induction occurred on medium containing 2, 4-D alone at 2 mg l−1 for ‘Bison’ and on medium containing 4 mg l−1 2, 4-D alone for ‘Bonilla’ big bluestem. For little bluestem, the highest percentage of embryogenic callus induction occurred on medium containing 3 mg l−1 2, 4-D plus 0.1 mg l−1 kinetin, suggesting that addition of KT is beneficial. Shoot regeneration took place on LS basal medium without any plant growth regulator for both species, although the addition of KT increased both regeneration frequency and the number of shoots produced per callus. Rooting of shoots reaching about 2 cm long occurred readily with or without α-naphthaleneacetic acid (NAA). Rooted plantlets were all successfully established in the soil.  相似文献   

4.
Efficient protocols were established for in vitro seed germination, neo-formation of secondary (2°) protocorms from primary (1°) protocorms and multiple shoot buds and protocorm-like body (PLB) induction from pseudo-stem segments of in vitro-raised seedlings of Cymbidium giganteum. Four nutrient media, namely Murashige and Skoog (MS), Phytamax (PM), Mitra et al. (M), and Knudson ‘C’ (KC) were evaluated for seed germination and early protocorm development. In addition, the effects of peptone, activated charcoal (AC) and two plant growth regulators [6-benzylaminopurine (BAP) and 2,4-dichlorophenoxyacetic acid (2,4-D)] were also studied. Both M and PM supplemented with 2.0 g l−1 peptone or 1.0 mg l−1 BAP resulted in ∼100% seed germination. Media supplemented with 2.0 g l−1 AC could effectively induce large protocorms (1.6 ± 0.1 mm in diameter). Neo-formation of 2° protocorms from 1° protocorms was achieved in liquid and agar-solidified PM medium fortified with different concentrations and combinations of auxins (α-naphthalene acetic acid (NAA) and 2,4-D) and cytokinins [BAP and kinetin (KN)]. The highest number of 2° protocorms was obtained in liquid medium (10.7 ± 0.9/1° protocorm) supplemented with 2.0 mg l−1 BAP + 1.0 mg l−1 NAA. Although protocorms proliferated profusely in liquid medium, these did not develop further unless transferred to agar-solidified medium within 6–8 weeks. Multiple shoot buds and PLBs were induced from pseudo-stem segments on agar-solidified PM medium fortified with different concentrations and combinations of BAP and NAA and the maximum number of PLBs (6.00 ± 0.20) was recorded when BAP and NAA were applied at 2.0 mg l−1 each. A solid root system was induced from PLBs and shoot buds when these were transferred to half-strength PM or M media fortified with 0.5 mg l−1 indole-3-acetic acid. Well-rooted plants were transferred to the greenhouse with 95% survival.  相似文献   

5.
An efficient method of repetitive somatic embryogenesis and plant regeneration was established in Coriandrum sativum L. Embryogenic callus was induced from cotyledon and hypocotyl segments on Murashige and Skoog (MS) medium with 4.52 μM 2,4-dichlorophenoxy acetic acid (2,4-D), upon subculturing on medium having same level of 2,4-D at an interval of 3 weeks developed somatic embryos, which progressed to cotyledonary stage through early developmental stages of somatic embryogenesis. The transfer of somatic embryos at an early cotyledonary and cotyledonary stage in clumps in succession to fresh 4.52 μM 2,4-D supplemented medium developed embryos in a cyclic manner. Upon transferal to embryogenic clumps (cotyledonary embryos) to modified MS medium (4 g l−1 KNO3, 0.29 g l−1 NH4NO3, 3 mg l−1 thiamine HCl, 0.5 mg l−1 pyridoxine HCl, and 5 mg l−1 nicotinic acid), the embryos irrespective of the cycles underwent maturation and germination. Germinating embryos transferred to half-strength MS medium favored healthy growth of plantlets. The system of recurrent somatic embryogenesis in coriander offers a system for genes transfer and also scale-up production of modified plants.  相似文献   

6.
Abiotic factors affect the induction of PLBs and callus in hybrid Cymbidium Twilight Moon ‘Day Light’. The initiation and proliferation of new PLBs and callus could be achieved on NAA and kinetin, supplemented at 0.1 mg l−1 each, respectively, both within 45–60 days. Bacto agar was found to be the most suitable solidifying agent for PLB induction, although a higher shoot fresh weight was obtained on Gelrite; a pH 5.3 was optimal while pH 4.5 caused 100% explant necrosis; coconut water, when supplied at 10–20% (v/v) resulted in a significant increase in the number of PLBs formed per PLB segment (23.1 versus 14.6 in controls) while a massive (almost four-fold) increase in fresh top weight occurred when PLB explants were placed in liquid culture, as a result of hyperhydricity; Fe-EDTA (1 mg l−1) and activated charcoal (1 g l−1) stimulated total fresh weight and PLB formation in the presence of PGRs; PLB formation decreased but total fresh shoot weight increased with the addition of niacin or myo-inositol, both vitamins. Dark-grown PLB-induced plants were etiolated and had longer internodes and higher fresh weight than light-grown control plants at 45 μmol m−2 s−1; at 15 μmol m−2 s−1 shoots were slightly etiolated, fragile, and PLB formation was scarce. RAPD and mtDNA analysis of all resultant PLBs, callus or plants showed them to be genetically identical, with comparable chlorophyll contents. Despite the detection of cytological variation between different plant parts, little variation resulted from abiotic factor treatment.  相似文献   

7.
“Fonio” (Digitaria exilis (L.) Stapf.) is a member of the grass family with excellent culinary and nutritional properties. In spite of its economic values, hardly has any improvement work been done. To enhance genetic improvement of this grain, plant regeneration protocol was developed using 8 cultivars. Stem segments of 5 mm long excised from 1 month-old seedlings germinated in vitro were cultured on 6 types of media for friable callus induction. Best result was obtained on Murashige and Skoog (MS) medium supplemented with 2 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 1 g l−1 casamino acid, where 91.3, 88.9 and 87.8% of the explants formed friable calli in cultivars ‘Kurelep’, ‘Churiwe’ and ‘Agyong’, respectively. Shoots appeared when friable calli were transferred to two regeneration media, i.e., MSBZ (MS medium + 0.022 mg l−1 2,4-D, 0 .22 mg l−1 6-benzylaminopurine (BA), 0.22 mg l−1 zeatin) and MSBG (MS medium + 0.5 mg l−1 BA, 0.1 mg l−1 gibberellic acid). The highest frequency of plant regeneration was attained on MSBG, with 91.7% of the friable calli forming shoots in cultivar “Churiwe”. Regenerated plants were rooted on hormone free MS medium. Flow cytometric analysis revealed 100% of the regenerants to be diploid. The protocol developed here can be used in the transformation of “Fonio” to increase the yield potential of this crop by incorporating characteristics such as disease resistance and stress resistance.  相似文献   

8.
Indiangrass [Sorghastrum nutans (L.) Nash.] is native to the North America and is an important component of the original tall grass prairie. It is also an important ornamental and forage grass. Recently, it has been proposed as an ideal biomass producer for cellulosic ethanol production. Genetic transformation is an important tool for introducing important agronomic traits into plants, but an efficient and reproducible in vitro regeneration protocol is a prerequisite for successful genetic transformation. In this report, we used mature caryopses as explants and tested the effect of various combinations of 2,4-dichlorophenoxyacetic acid (2,4-D) (1–5) and kinetin (KT) (0, 0.1, and 0.2) on embryogenic callus induction using LS basal medium. Caryopses cultured on media supplemented with 2,4-D alone generally outperformed those cultured on media supplemented with both 2,4-D and kinetin for embryogenic callus induction. The best treatment is LS basal medium supplemented with 3 mg l−1 2,4-D. LS basal medium supplemented with KT of 0, 0.5, 1, 2 or 5 mg l−1 were tested for regeneration efficiency which was shown to increase as the KT concentration increased. The quality of the shoots produced on the medium containing KT at 5 mg l−1, which produced the highest regeneration frequency appeared to be lower as leaves become vitrified. Shoots were moved to a rooting medium containing either 0 or 0.1 mg l−1 α-naphthaleneacetic acid (NAA). Rooted plantlets were then transferred to soil-containing pots and were placed in a mist room for 1 week before they are transferred to a normal greenhouse where they all survived. The reported regeneration protocol is very efficient and highly reproducible in spite of the heterogeneous nature of the tested cultivar; thus it should be suitable for genetic transformation.  相似文献   

9.
Present study demonstrates the effect of sucrose and ABA on germination of encapsulated somatic embryos of guava (Psidium guajava L.). Sucrose and ABA at different concentrations were also evaluated for their effects on maturation and germination of somatic embryos. Mature somatic embryos developed on MS medium containing high concentration of sucrose (10%) or ABA (1.0 mg l−1) showed inhibition in germination if they continued to be in same medium for 4 weeks. With increasing concentrations of sucrose (3–9%) or ABA (0.01–1.0 mg l−1) in medium, percent germination of encapsulated somatic embryos decreased significantly. Encapsulated somatic embryos after storage on MS medium supplemented with 9% sucrose or 1 mg l−1 ABA for different duration (0–60 days) germinated when they were transferred to medium containing 3% sucrose. About 20.8% and 37.5% encapsulated somatic embryos germinated after storage on ABA (1 mg l−1) or sucrose (9%) for 60 days, respectively. Temporarily suppression in germination of encapsulated somatic embryos by high concentration of sucrose or ABA may be important for short-term conservation of elite genotype of guava.  相似文献   

10.
Lychnis (Caryophyllaceae) consists of about 30 species distributed throughout the temperate regions of the Northern Hemisphere, from East Asia to Europe. Many Lychnis spp. have high ornamental value and cultivated as pot or garden plants. In the present study, in vitro chromosome doubling of several Lychnis spp. was examined in order to widen their variability in horticultural traits. Initially effect of various spindle toxin treatments [100, 500 or 1000 mg l−1 colchicine (COL), 10, 20 or 50 mg l−1 oryzalin (ORY), or 1, 5, 10 mg l−1 amiprophos-methyl (APM)] of nodal segments of a triploid genotype of L. senno (3x) was investigated on survival of nodal segments and chromosome doubling in nodal segment-derived plantlets. Significantly higher percentage (75.0%) of surviving segments after spindle toxin treatment was obtained in 10 mg l−1 ORY treatment. Flow cytometry (FCM) analysis of leaf tissues showed that 9.4–13.8% of plantlets, which were derived from 10 to 20 mg l−1 ORY, or 5 mg l−1 APM treatments, were hexaploid (6x) or ploidy chimera (3x + 6x, 4x + 6x, 5x + 6x, 3x + 4x + 6x). The results obtained by FCM analysis were confirmed by chromosome observation in root tip cells. Thus 10 mg l−1 ORY treatment of nodal segments is suitable for in vitro chromosome doubling of triploid L. senno. Efficient chromosome doubling was also achieved in diploid L. fulgens (2x) and L. sieboldii (2x) by treating nodal segments with 10 mg l−1 ORY: 68.9–88.7% of nodal segments survived after ORY treatment, and 24.7–26.5% of plantlets derived from ORY-treated nodal segments were tetraploid (4x) or ploidy chimera (2x + 4x) in both species. These results indicate that the in vitro chromosome doubling method established for triploid L. senno may be applicable to a wide range of Lychnis spp. Tetraploid L. fulgens and L. sieboldii showed a compact plant form, and had thick stems and deep green leaves compared with the diploid mother plants. On the other hand, hexaploid L. senno showed very poor growth and died before flowering.  相似文献   

11.
An in vitro plant regeneration protocol for pansy (Viola wittrockiana) cultivar ‘Caidie’ from petioles was established as following: callus induction on a half-strength MS medium supplemented with 0.45 μmol l−1 2,4-d plus 8.9 μmol l−1 BA, callus subculture on medium F (1/2MS with 4.5 μmol l−1 2,4-d, 2.7 μmol l−1 NAA and 0.44 μmol l−1 BA) and then on medium T (1/2MS with 4.5 μmol l−1 2,4-d, 2.7 μmol l−1 NAA and 2.2 μmol l−1 BA), shoot regeneration on medium D3 (MS media supplemented with 2.9 μmol l−1GA3, 23.6 μmol l−1 AgNO3, 0.02% active charcoal and 4.5 μmol l−1 TDZ), shoot multiplication on medium M (half-strength MS medium containing NAA 1.1 μmol l−1, TDZ 9.1 μmol l−1 and GA3 8.7 μmol l−1), and then shoot elongation and rooting on medium R (MS medium supplemented with 1.1 μmol l−1 NAA and 1.1 μmol l−1 BA). Subculture on appropriate medium was found to be important for successful shoot regeneration.  相似文献   

12.
The proliferation of embryogenic suspension culture in two cultivars (Jihel and Bousthami Noir) of Phoenix dactylifera L. was tested on liquid media with or without 2,4-d and with different glutamine concentrations (3.35 × 10−4, 6.7 × 10−4 and 13.4 × 10−4 M). The liquid medium with 0.1 mg l−1 2,4-d and 6.7 × 10−4 M glutamine has clearly improved the proliferation of somatic embryos. In fact, when glutamine concentration increased from 3.35 × 10−4 to 6.7 × 10−4 M, the yield of somatic embryos increased from 14 to 56 embryos per 100 ml of culture medium for “Jihel” cultivar and 25–71 embryos per 100 ml of culture medium for “Bousthami Noir” cultivar. In contrast, increasing glutamine concentration from 6.7 × 10−4 to 13.4 × 10−4 M, the embryos yield was negligible. Based on biochemical analysis, the highest accumulation of proteins and sugars was obtained in liquid medium with 0.1 mg l−1 2,4-d and 6.7 × 10−4 M glutamine (118 and 91 mg of proteins g−1 DW, respectively, for “Jihel” and “Bousthami Noir” cultivars; 194 mg of sugars g−1 DW for “Jihel” cultivar and 182 mg of sugars g−1 DW for “Bousthami Noir” cultivar). In addition, the supply of 0.05 mg l−1 BAP on the germination medium could be useful in terms of germination percentage of somatic embryos. When BAP concentration increased from 0.05 to 0.2 mg l−1, the germination percentage of somatic embryos decreased from 14.2 to 4.9%, while secondary embryogenesis increased from 26.4 to 45.2%.  相似文献   

13.
Ornithogalum ulophyllum Hand.-Mazz. with beautiful white flowers is an important medicinal and ornamental plant of the Middle Eastern countries and need exploitation for commercial propagation. The study reports in vitro mass proliferation of bulblets achieved from twin scales and “in vitro regenerated bulblet” explants on MS medium supplemented with various concentrations of BAP–NAA. The best regeneration on twin scales and “in vitro regenerated bulblets” was obtained on MS medium containing 2 mg l−1 BAP–0.5 mg l−1 NAA and 2 mg l−1BAP–1 mg l−1 NAA, respectively. However, bulb scales seemed to be more potent for bulblet regeneration. A large number of the developing bulblets rooted on the regeneration medium. Remaining non-rooting bulblets were rooted on MS medium containing 1 mg l−1 NAA. All plants were acclimatized in the environmental chamber for 4 weeks and were transferred to the greenhouse for flowering. Regenerated bulblets developed into morphologically normal plants.  相似文献   

14.
The genus Ptilotus has immense potential for ornamental horticulture but its commercial development has been hindered by propagation limitations. Poor seed quality and germination are reported. Cutting propagation is limited by cutting supply as the juvenile phase of Ptilotus is short. Micropropagation has been used in an attempt to overcome these difficulties but explants become floral in vitro and this causes plantlets to elongate. Ethephon has been used to control flowering of stock plants of many ornamental species. This study investigated the effect of ethephon applied to young (3-week-old, deflasked from tissue culture) and mature (1-year-old) Ptilotus plants in a greenhouse. A system of applying gaseous ethylene at 0, 100, 200 and 300 mg l−1 to the headspace of in vitro plantlets in glass jars was developed and the response of in vitro plantlets to ethylene studied. One-year-old Ptilotus plants were treated with 500 mg l−1 ethephon 2 days before pruning or 1 or 2 weeks after pruning. Ethephon application 2 days before pruning decreased the number of inflorescences and increased the number of shoots (compared to the control) but was phytotoxic. Ethephon applications of 150 or 300 mg l−1 applied weekly or fortnightly to 3-week-old plants deflasked from tissue culture reduced plant height and number of inflorescences and at low concentrations increased the number of new shoots. A fortnightly application at 150 mg l−1 is recommended. Previous reports on the effects of ethylene on inflorescence production on plantlets in vitro are limited. Our study showed that exposure of in vitro plantlets of P. nobilis to ethylene gas at 100 mg l−1 for 1 h significantly increased the number of shoots and plant height but this did not occur for plantlets of P. spicatus. Plantlets of P. spicatus exposed to transient ethylene at 200 and 300 mg l−1 showed significantly greater rooting (52.4%) than the control (13.6%).  相似文献   

15.
Methods to regenerate whole plants from mature leaf explants of Pelargonium rapaceum (L.) L’Hérit were established. To optimize shoot induction, leaf explants were cultured on media containing different types and combinations of plant growth regulators. Growth was initiated within 17–24 days culture, and included callus formation, and root or shoot organogenesis ranging from 20 to 100% regeneration. Shoots were induced only when explants were cultivated on MS medium containing a combination of NAA and kinetin, NAA and BAP, IAA and Kinetin, or IAA and BAP. On media containing NAA and BAP, dark incubation was critical for efficient direct shoot regeneration from explants. Direct shoot formation and the highest number of shoots per explant (17.6) were obtained from leaf explants cultured in the dark for 30 days on MS medium containing 0.1 mg l−1 NAA and 0.1 mg l−1 BAP. Shoots cultured on MS medium containing 0.1 mg l−1 NAA formed tuberous roots with microtubers within 42 days. Healthy regenerated plants were acclimated and transferred to a greenhouse.  相似文献   

16.
Pogonatherum paniceum (Lam.) Hack. is a rock plant with good potential for vegetative recovery on naked lands. A high frequency in vitro regeneration system was developed for P. paniceum. Calli were induced from explants of mature seeds, seedlings, young leaves, and stem segments on Murashige and Skoog (MS) medium supplemented with 1.0 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D), 2.0 mg L−1 α-naphthalene acetic acid (NAA) and 0.2 mg L−1 6-benzylaminopurine (BAP). High induction rates (59.57%) and regeneration rates (100%) were obtained from mature seed explants; calli were sub-cultured for over 2 years and still retained a high regenerative capacity. One seed explant resulted in 69,997 plants in 1 year. Shoot buds derived from calli were used for encapsulation in liquid MS medium containing 3% sucrose and two different alginate matrices (3% sodium alginate (w/v) + MS medium containing 3% sucrose and 3% sodium alginate + 1% activated carbon (w/v) + MS medium containing 3% sucrose) with a 20-min exposure to 2% CaCl2 and 0.3% bavistin (w/v). The capsule with 3.0% sodium alginate (w/v) and 1% activated carbon (w/v) showed a higher conversion rate (61.58%) and stronger plantlets under non-aseptic conditions. These systems are useful for the rapid clonal propagation and dissemination of artificial seed material of P. paniceum for eco-recovery.  相似文献   

17.
The regenerability of three ornamental species—Lysimachia christinae, Lysimachia rubinervis and Lysimachia nummularia ‘Aurea’, were investigated using in vitro leaves and shoot tips. 6-Benzylaminopurine (BAP) and α-naphthalene acetic acid (NAA) added to Murashige and Skoog (MS) medium were tested for their effect on organogenesis. On the medium, shoot regeneration occurred directly without callus formation. In these species, L. christinae developed the highest regeneration rate and numbers of shoots/explant from shoot tips (100%, 12.25) and leaf bases (100%, 13.01) on the MS medium containing 3.0 mg l−1 BAP and 0.1 mg l−1 NAA. For L. rubinervis, the highest shoot induction rate and number of shoots/explant were obtained from shoot tip (100%, 16.87–17.20) on the MS medium with 0.1 mg l−1 NAA and 3.0–5.0 mg l−1 BAP. L. nummularia ‘Aurea’, however, showed the highest regeneration rate and number of shoots/explant (100%, 12.73) from leaf bases on MS medium supplemented with 1.0 mg l−1 BAP and 0.1 mg l−1 NAA. All in vitro shoots rooted well on half macronutrient MS medium containing 0.1 mg l−1 NAA. After acclimatization, transplanted plantlets grew normally and flowered in the field.  相似文献   

18.
High frequency and direct (without callus) plant regeneration was achieved from whole leaf explants of thornless blackberry (Rubus hybrid) cv. Black Satin (EC No. 381258; PI No. 553272) in vitro. Leaf blade explants from 1-, 3- and 5-month-old mother cultures were cultured on Murashige and Skoog (MS) medium with thidiazuron (TDZ), N6-benzylaminopurine (BAP), indol-3-butyric acid (IBA) and α-naphthalene acetic acid (NAA), alone or in combination. Three-month explants cultured on 0.02 mg l−1 TDZ produced a high regeneration frequency (91.7%) and the most shoots/leaf explant (17.3). The shoot primordia developed within 3 weeks from the point of detachment of the petiole from the leaf blade. The age of the explant source significantly affected the shoot regeneration potential of the leaf explants. Leaves excised from 3-month-old in vitro-cultured shoots performed better than those from 1- and 5-month-old shoots. Shoots rooted best on half-strength MS basal medium with 0.5 mg l−1 IBA and 90% of the plantlets survived acclimatization. The regenerated plantlets were morphologically similar to the mother plants.  相似文献   

19.
Inter-section hybrids were obtained in the reciprocal crosses between Primula filchnerae (2n = 2x = 24) of Sect. Pinnatae and P. sinensis ‘Fanfare’ (2n = 2x = 24) of Sect. Auganthus by rescuing ovules on half-strength (1/2) Murashige and Skoog's (MS) medium supplemented with 50 g l−1 sucrose, 2.5 g l−1 gellan gum, 0.1 mg l−1 α-naphthaleneacetic acid (NAA), 0.1 mg l−1 6-benzyladenine (BA) and 50 mg l−1 gibberellic acid (GA3). In ovule culture, germination occurred with radicle elongation but no plumule was observed. The radicle kept on the initial medium showed root proliferation with callus formation. When the calluses were transferred to (1/2)MS media containing 30 g l−1 sucrose and 3 g l−1 gellan gum, without plant growth regulators (PGRs) or with 1 mg l−1 zeatin and 0.1 mg l−1 NAA, plantlets were regenerated. The plants thus obtained were confirmed to be hybrids through flow cytometry (FCM) and random amplified polymorphic DNA (RAPD) analyses. The hybrid obtained when P. filchnerae was used as the maternal parent was diploid, whereas hexaploid hybrid was obtained when using P. sinensis as the maternal parent. The hexaploid hybrid might be produced through chromosome doubling of a triploid originated from the fertilization of P. sinensis with unreduced pollen of P. filchnerae.  相似文献   

20.
Seedless grapevine cultivars (Vitis vinifera L.) are widely grown in Europe, America and Asia. Abortion of zygotic embryos in seedless grapes largely limits the efficiency of breeding of seedless cultivars through genetic crossing. The present study was designed to investigate effects of exogenously applied plant growth regulators (PGRs) to the grapevines in field condition on ovule and subsequent embryo rescue of seedless grapes of small seed traces. First experiment was performed by measuring ovules weight, proportion of each category ovules in maturity and embryo development in vitro of seedless grape cv. Centennial Seedless, Thompson Seedless and Crimson Seedless sprayed by chlormequat (CCC), benzyladenine (BA), ethephon (CEPA) and putrescine (Put). The effects of different application concentration and date of CCC were further evaluated in Centennial Seedless in later experiment. The results showed that exogenous application of all PGRs did not affect the total number of ovules per berries in maturity. CCC increased the ovules weight and proportion of ovules >4 mm in length of three varieties in maturity. The effects of two application times of PGRs on weight of berries and ovules and proportion of each category ovules in maturity were not significantly different. In the proceeding of embryo rescue, CCC at 100 and 1000 mg l−1, BA at 100 mg l−1 and Put at 20 mg l−1 increased the percentage of developed embryos of Centennial Seedless and Thompson Seedless. The results showed that the size of ovules excised for embryo rescue significantly affected embryo formation and plant regeneration. The percentage of embryos formation in ovules >4 mm in length was significantly more than in ovules 2–4 mm in length, no embryo was found in ovules <2 mm in length. Exogenous application of CCC at 100–500 mg l−1 significantly increased percentage of ovules >2 mm in length by 80.0–82.7% in Centennial Seedless, therefore improving embryo formation. The statistical correlation was found between the proportion of ovules >2 mm and embryo formation (r = 0.92) in Centennial Seedless. Among the different spraying time in Centennial Seedless, CCC applied 14 days before bloom produced significantly more ovules >2 mm in length and embryos formation.  相似文献   

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