Comparison of NAA and 2,4-D induced somatic embryogenesis in Cassava

NAA and 2,4-D were compared for their ability to induce somatic embryogenesis in cassava (Manihot esculenta Crantz). In all seven cultivars tested, only 2,4-D had the capacity to induce primary somatic embryos from leaf explants, however, both NAA and 2,4-D were capable of inducing secondary somatic embryos. More secondary somatic embryos were formed in NAA than in 2,4-D medium. Furthermore, the maturation period for secondary somatic embryos was shorter in NAA medium than in 2,4-D medium. In some cultivars, repeated subculture of secondary somatic embryos in NAA medium resulted in a gradual shift from somatic embryogenesis to adventitious root formation. This shift could be stopped and reversed by subculture of the material in 2,4-D medium. In NAA medium the most secondary somatic embryos were formed when they were subcultured every 15 days whereas in 2,4-D a 20 day subculture interval was optimal. Subculture of secondary somatic embryos at a high inoculum density (>1.5 g jar⁻¹) in NAA medium did not result in the formation of secondary somatic embryos, whereas in 2,4-D it lead to the formation of globular secondary somatic embryos. With 2,4-D the newly induced secondary somatic embryos were connected vertically to the explant and with NAA medium horizontally. For all cultivars tested, desiccation stimulated normal germination of NAA-induced somatic embryos. However, the desiccated, secondary somatic embryos required a medium supplemented with BA for high frequency germination. The concentration of BA needed for high frequency germination was higher when the desiccated secondary somatic embryos were cultured in light instead of dark. In only one cultivar desiccation enhanced germination of 2,4-D induced secondary somatic embryos and in three other cultivars it stimulated only root formation. This revised version was published online in June 2006 with corrections to the Cover Date.     

Enhanced Maturation and Desiccation Tolerance of White Spruce [Picea glauca (Moench) Voss] Somatic Embryos: Effects of a Non-plasmolysing Water Stress and Abscisic Acid

A non-plasmolysing moisture stress effected by polyethyleneglycol (PEG) was beneficial when applied to maturing white spruce(Picea glauca) somatic embryos for the following reasons. Anosmotic treatment of 5.0–7.5% PEG stimulated a threefoldincrease in the maturation frequency. The osmotically treatedsomatic embryos displayed higher dry weights and lower moisturecontents than the controls, indicating a greater accumulationof storage reserves. Moisture contents of mature, osmotically-treated,hydrated somatic embryos were 40–45%, in contrast to 57%for the non-osmotically treated controls. Desiccation was achievedby placing the somatic embryos in a range of relative-humidityenvironments. No clear trend for the effect of PEG on survivalof desiccated somatic embryos was observed; mean survival valuesranged from 34 to 62% when somatic embryos from all osmotictreatments were desiccated for 14 d at 81% relative humidity.Following this desiccation treatment, somatic embryos from allosmotic concentrations had moisture contents of 26–31%,similar to the 32% recorded for unimbibed zygotic embryos. Afterimbibition, moisture contents for these zygotic and somaticembryos were in the order of 60%. Somatic embryos matured withPEG remained quiescent during desiccation due to their low initialmoisture contents, and gave rise to plantlets of normal appearance.Gradual desiccation of the somatic embryos directly followingmaturation with abscisic acid (ABA) was crucial to survivalduring desiccation. A plasmolysing water stress effected bysucrose at osmotic potentials similar to PEG was detrimentalto somatic embryo maturation, thereby emphasizing the importanceof the choice of osmoticum. Desiccation, maturation, osmotic potential, Picea glauca, polyethylene glycol, somatic embryo, water stress, white spruce     

Effects of proliferation,maturation, and desiccation methods on conversion of soybean somatic embryos

Summary Cermination of soybean [Glycine max (L.) Merrill] somatic embryos and conversion to whole plants are generally low. This study was conducted to investigate the effects of proliferation, maturation, and desiccation methods on conversion of soybean somatic embryos to plants. Soybean cv. Jack somatic embryos, proliferated on a solid medium containing 90.5 μM (20 mgl⁻¹) 2.4-dichlorophenoxyacetic acid (2.4-D) (MSD20), showed a regeneration rate signficantly higher than those proliferated in a liquid medium containing 45.25 μM (10mgl⁻¹) 2,4-D (FN Lite). When a liquid medium without 2,4-D and B5 vitamins (FN Superlite) was used for maturation, the duration of time necessary for embryo development could be shortened by more than a month compared to maturation on a standard solid medium (MSM6AC). An air-drying method, in which somatic embryos were desiccated in an empty sealed Petri dish for 3–5d, gave rise to the best germination efficiency among the four desiccation methods tested: fast, slow, air, and KCl methods. The final percentage of moisture seems important since embyros over-dried by the fast and slow methods did not convert well into plants.     

Regulation of starch and protein accumulation in alfalfa (Medicago sativa L.) somatic embryos

Compared to seeds, somatic embryos accumulated relatively low levels and different types of storage carbohydrates. The regulation of starch accumulation was studied to determine its effects on desiccation tolerance and vigor of dry somatic embryos. Somatic embryos of Medicago sativa are routinely matured through three phases: 7 days of development; 10 days of phase I maturation, a rapid growth phase; and 10 days of phase II maturation, a phase leading to the acquisition of desiccation tolerance. The control of starch deposition was investigated in alfalfa somatic embryos by manipulating the composition of the phase I maturation medium with different levels of sucrose, abscisic acid, glutamine and different types of carbohydrates and amino acids. After phase II maturation, mature somatic embryos were collected for desiccation and subsequent conversion, or for biochemical analyses. Starch deposition occurred primarily during phase I maturation, and variations in the composition of this medium influenced embryo quality, storage protein and starch accumulation. A factorial experiment with two levels of glutamine × three levels of sucrose showed that increasing the sucrose concentration from 30 to 80 g/l increased embryo size and starch content, but had minimal effect on accumulation of storage proteins; glutamine also increased embryo size, but decreased starch content and increased accumulation of the high salt soluble S-2 (medicagin) storage proteins. ABA did not influence any of the parameters tested when included in phase I maturation at concentration up to 10 μM. Replicating sucrose with maltose, glucose, or glucose and fructose did not alter embryo size or starch accumulation (mg/g fresh weight), but replacement with fructose alone reduced embryo size, and replacement with glucose alone reduced germination. Suplementation with the amino acids, asparagine, aspartic acid and glutamine increased seedling vigor, but decreased the starch content of embryos. The data indicate that starch accumulation in somatic embryos is regulated by the relative availability of carbon versus nitrogen nutrients in the maturation medium. The quality of mature somatic embryos, determined by the rate of seedling development (conversion and vigor), correlated with embryo size, storage protein and free amino acid but not with starch. Therefore, further improvements in the quality of somatic embryo may be achieved through manipulation of the maturation medium in order to increase storage protein, but not starch deposition.     

Improving tolerance of somatic emrbyos of Picea glauca to flash desiccation with a cold treatment (desiccation after cold acclimation)

Summary Controlled mild desiccation of mature white spruce somatic embryos prior to germination improves the quality of the germinated embryos. More severe desiccation results in increased injury and death but is desirable for long-term storage of embryos and production of desiccated artificial seed. A method was developed to improve desiccation tolerance in somatic embryos using a temperature treatment. Culture plates with embryos at four stages of development were subjected to temperatures of 1, 5, 10, or 20°C for periods of 0, 1, 2, 4, or 8 wk duration. After the temperature treatment, the embryos were harvested and air-dried for 2h under a laminar flow hood. Dried embryos were placed directly on germination medium and the quality of the germinants was assessed after 4 wk. The initial maturation stage of the embryo and the temperature and duration of the treatment had a significant effect on the quality of the germinants. Most treatments caused marked differential survival of organs. The optimal response was obtained with embryos that had been grown for 51 d (cotyledonary stage) on maturation medium and that were subsequently exposed to a temperature of 5°C for 8 wk prior to air drying. This treatment produced 58% undamaged germinants with normal cotyledons, hypocotyls, and roots. Only 1% of the untreated air-dried embryos germinated normally.     

Manipulating the desiccation tolerance and vigor of dry somatic embryos of Medicago sativa L. with sucrose,heat shock and abscisic acid

Summary The effects of sucrose concentration in the maturation medium in combination with a heat shock treatment at 36°C were investigated in an attempt to improve the vigor of seedlings grown from dry somatic embryos of alfalfa (Medicago sativa L.). Callus was formed from petiole expiants and dispersed in liquid suspension medium in the presence of 5 M 2,4-D. The cell suspension was sieved to synchronize embryo development. The 200 – 500 m fraction was plated on embryo development medium without 2,4-D, grown for 14 days, and transferred to maturation medium. With 3% sucrose in the maturation medium, the somatic embryos germinated precociously and were unable to survive desiccation. At higher sucrose concentrations, germination was delayed and the embryos continued to accumulate dry matter. After 13 days on 6% sucrose medium (27 days after sieving), the somatic embryos were tolerant of drying to 12% moisture without exposure to exogenous ABA. Heat shock, which presumably stimulates endogenous ABA synthesis, improved the desiccation tolerance of somatic embryos if applied prior to day 27 after sieving, but its effects were minimal after day 27. High sucrose concentrations up to 9% in the maturation medium were optimal during the first 8 days on maturation medium (days 14 to 22 after sieving), but a lower concentration (6%) was optimal during the later stages of embryo maturation (days 22 to 30 after sieving). The inclusion of 10^–5 M ABA in the maturation medium with 6% sucrose further improved embryo quality if applied approximately 20 days after sieving.     

Peroxidase activity of desiccation-tolerant loblolly pine somatic embryos

Summary Desiccation tolerance can be induced by culturing somatic embryos of loblolly pine (Pinus taeda L.) on medium supplemented with 50 μM abscisic acid (ABA) and/or 8.5% polyethylene glycol (PEG₆₀₀₀). Scanning electron microscopy of desiccated somatic embryos showed that the size and external morphology of the desiccation-tolerant somatic embryos recovered to the pre-desiccation state within 24–36 h, whereas the non-desiccation-tolerant somatic embryos did not recover and remained shriveled, after rehydration. Peroxidase activity of desiccated somatic embryos increased sharply after 1 d of desiccation treatment at 87% relative humidity (RH), and desiccation-tolerant somatic embryos had higher peroxidase activity compared to sensitive somatic embryos. Higher peroxidase activity of desiccation-tolerant somatic embryos may have allowed them to catalyze the reduction of H₂O₂ produced by drought stress, and protected them from oxidative damage.     

Improvement of somatic embryogenesis and plant recovery in cassava

Methods for improving the efficiency of plant recovery from somatic embryos of cassava (Manihot esculenta Crantz) were investigated by optimizing the maturation regime and incorporating a desiccation stage prior to inducing germination. Somatic embryos were induced from young leaf lobes of in vitro grown shoots of cassava on Murashige and Skoog medium with 2,4-dichlorophenoxy acetic acid. After 15 to 20 days of culture on induction medium, the somatic embryos were transferred to a hormone free medium supplemented with activated charcoal. In another 18 days mature somatic embryos became distinctly bipolar and easily separable as individual units and were cultured on half MS medium for further development. Subsequent desiccation of bipolar somatic embryos resulted in 92% germination and 83% complete plant regeneration. The plants were characterized by synchronized development of shoot and root axes. Of the non-desiccated somatic embryos, only 10% germinated and 2% regenerated plants. Starting from leaf lobes, transplantable plantlets were derived from primary somatic embryos within 70 to 80 days.Abbreviations 2,4-D 2,4 dichlorophenoxyacetic acid - BA Benzyl aminopurine - GA Giberellic acid - MS Murashige and Skoog - NAA Naphthalene acetic acid     

Effect of exogenous ABA on somatic embryo maturation and germination in Persian walnut (<Emphasis Type="Italic">Juglans regia</Emphasis> L.)

Low efficiency of embryo maturation, germination and conversion to plantlets is a major problem in many species including Persian walnut. We studied the effects of abscisic acid (ABA) and sucrose, on the maturation and germination of Persian walnut (Juglans regia) somatic embryos. Individual globular somatic embryos were grown on a maturation medium supplemented with different combinations of ABA and sucrose for ca. 1 month, until shoot meristems and radicles had developed. White and opaque embryos in late cotyledonary stage were subjected to desiccation after the culture period on maturation media. The number of germinated somatic embryos was influenced by the concentrations of ABA in the maturation medium. The best treatment for germination, in which both shoot and root were developed contained 2 mg l⁻¹ ABA and resulted in 41% conversion of embryos into plantlets. Regeneration was reduced at higher levels of ABA. While ABA always reduced the rate of secondary embryogenesis, treatments containing 4.0% sucrose significantly increased the number of secondary embryos. On the other hand, sucrose had little influence on maturation. Normal and abnormal embryos were verified anatomically.     

Manipulation of conditions for the culture of somatic embryos of white spruce for improved triacylglycerol biosynthesis and desiccation tolerance

In order to enhance post-germinative vigour, somatic embryos of Picea glauca (Moench) Voss. were matured under in-vitro conditions that stimulated triacylglycerol (TAG) biosynthesis. In P. glauca seeds over 90% of the TAG was stored within the megagametophyte, and isolated zygotic embryos contained twice the amount of TAG of somatic embryos cultured for four weeks on basal medium containing 16 M abscisic acid (ABA). Polyethylene glycol-4000 (PEG) as a non-permeating osmoticum with ABA promoted TAG biosynthesis by somatic embryos and sustained maturation throughout an eight-week culture period. Treatments that promoted TAG biosynthesis also prevented precocious germination and promoted desiccation tolerance. Thus, the optimal culture conditions for maturation, desiccation survival, and plantlet regeneration were 16–24 M ABA and 7.5% PEG for eight weeks, followed by desiccation. Under these conditions the levels of TAG per somatic embryo were raised ninefold to about five times the zygotic-embryo level, and the TAG fatty-acid composition became similar to that of zygotic embryos. A study of sectioned material, using light and transmission electron microscopy, showed that the structure and distribution of lipid bodies within these somatic embryos and the degree of embryo development were similar to mature zygotic embryos. Up to 81% of the desiccated somatic embryos regenerated to plantlets during which time the TAG was utilised in a manner similar to zygotic seedlings.Abbreviations ABA abscisic acid - PEG polyethylene glycol - TAG triacylglycerol - TL total lipid - TEM transmission electron microscopy Plant Research Centre contribution No. 1383We are grateful to Dawn Moore and Ken Stanley for technical assistance, and thank Pat Rennie for the electron microscopy. We acknowledge financial support through an NSERC/Forestry Canada/Weyerhaeuser Canada Ltd (Prince Albert, Sask.) research partnership programme.     

Direct somatic embryogenesis and synthetic seed production from<Emphasis Type="Italic"> Paulownia elongata</Emphasis>

We have developed a reproducible system for efficient direct somatic embryogenesis from leaf and internodal explants of Paulownia elongata. The somatic embryos obtained were subsequently encapsulated as single embryos to produce synthetic seeds. Several plant growth regulators [6-benzylaminopurine, indole-3-acetic acid, -naphthaleneacetic acid, kinetin and thidiazuron (TDZ)] alone or in combination were tested for their capacity to induce somatic embryogenesis. The highest induction frequencies of somatic embryos were obtained on Murashige and Skoog (MS) medium supplemented with 3% sucrose, 0.6% Phytagel, 500 mg l^-1 casein hydrolysate and 10 mg l^-1 TDZ (medium MS10). Somatic embryos were induced from leaf (69.8%) and internode (58.5%) explants on MS10 medium after 7 days. Subsequent withdrawal of TDZ from the induction medium resulted in the maturation and growth of the embryos into plantlets on MS basal media. The maturation frequency of somatic embryos from leaf and internodal explants was 50.8% and 45.8%, respectively. Subculturing of mature embryos led to their germination on the same medium with a germination frequency of 50.1% and 29.8% from leaf and internode explants, respectively. Somatic embryos obtained directly on leaf explants were used for encapsulation in liquid MS medium containing different concentrations of sodium alginate with a 30-min exposure to 50 mM CaCl₂. A 3% sodium alginate concentration provided a uniform encapsulation of the embryos with survival and germination frequencies of 73.7% and 53.3%, respectively. Storage at 4°C for 30 days or 60 days significantly reduced the survival and complete germination frequencies of both encapsulated and non-encapsulated embryos relative to those of non-stored somatic embryos. However, the survival and germination rates of encapsulated embryos increased following storage at 4°C. After 30 days or 60 days of storage, the survival rates of encapsulated embryos were 67.8% and 53.5% and the germination frequencies were 43.2% and 32.4%, respectively. These systems could be useful for the rapid clonal propagation and dissemination of synthetic seed material of Paulownia elongata.Abbreviations BAP 6-Benzylaminopurine - IAA Indole-3-acetic acid - NAA -Naphthaleneacetic acid - TDZ ThidiazuronCommunicated by H. Lörz     

Polyethylene glycol promotes maturation but inhibits further development of Picea abies somatic embryos

A combined application of abscisic acid (ABA) and high molecular mass osmoticum, polyethylene glycol (PEG) has become a routine method for stimulating somatic embryo maturation in some genera of Coniferales. The goals of the present study were to clarify how the PEG 4000-attributed low osmotic potential (ψ_s) of the maturation medium affects the yield and morphology of mature somatic embryos as well as subsequent developmental processes during germination and ex vitro plantlet growth in different genotypes of Picea abies belonging to 3 full sib seed families. Despite high within- and among-family variation, a stimulatory effect of 7.5% PEG (ψ_s=?0.645 MPa) on somatic embryo maturation was recorded for 13 out of 17 cell lines (F= 2.83, P= 0.1). PEG-treated somatic embryos were more dehydrated than embryos matured in the absence of PEG. Subsequently, embryos were partially desiccated using a high relative humidity treatment (HRH-treatment). The dynamics of embryo water content (WC) during HRH-treatment differed between embryos developed on maturation medium for 5 or 7 weeks. These two patterns remained unchanged irrespective of the ψ_s of the maturation medium. In 5-week somatic embryos, the WC decreased to the lowest level (in the range 25-35%) within the first 8 days of HRH-treatment and was not further substantially changed. Seven-week embryos also lost water within 8 to 16 days (decrease to 15-25% WC), but this drop was followed by rehydration of embryonic tissues by 24th day of HRH-treatment up to nearly the initial WC. Thus, 7-week embryos experienced both desiccation and slow imbibition in the course of the 24-day HRH-treatment. This could account for their increased germinability compared to 5-week somatic embryos found in the present study. Addition of 7.5% PEG to the maturation medium significantly inhibited somatic embryo germination for the vast majority of genotypes (F= 7.35; P= 0.01). Moreover, even after ex vitro transfer, both radicle elongation and lateral root formation were substantially suppressed (F= 3.8; P= 0.03) in those plantlets produced from PEG-treated somatic embryos. Alterations both in the organization of the root meristem and in the structure of the root cap were found by histomorphological analysis of PEG-treated somatic embryos. All those embryos possessed massive root caps with numerous intercellular spaces in the pericolumn tissue. Cells of the quiescent center exhibited clear symptoms of degradation manifested in shrinkage and collapse of the protoplasm. In addition, PEG-treated embryos were of smaller size compared to embryos matured without osmoticum. When grown in artificial substrate (up to 5 months) the PEG-induced inhibitory post-effect gradually decreased. At this stage, the duration of maturation was the only factor separating plantlets into slow- and fast-growing categories. Somatic embryos matured for 5 weeks produced plantlets twice the size of those produced by 7-week embryos (F= 37.8; P < 0.0001). This trend did not depend on ψ_s of the maturation medium, nor on the genotype.     

Dormancy induction of somatic embryos of Siberian ginseng by high sucrose concentrations enhances the conservation of hydrated artificial seeds and dehydration resistance

. In most plants, somatic embryos tend to germinate prematurely, a process that is detrimental to controlled plant production and the conservation of artificial seeds. We investigated the dormancy characteristics of Siberian ginseng somatic embryos induced simply by a high sucrose treatment, a treatment that enables the long-term conservation of artificial seeds following encapsulation and provides embryos with an enhanced resistance to dehydration. Early-cotyledonary stage somatic embryos were mass-produced by means of bioreactor culture. These embryos were then plated on medium supplemented with various levels of sucrose (1%, 3%, 6% or 9%) and allowed to mature. Subsequent germination of these embryos following the maturation period depended significantly on the sucrose level. At concentrations of 9% sucrose, none of the somatic embryos germinated after maturation, and none were recovered after being transferred to half-strength MS medium containing 2% sucrose. Gibberellic acid treatment was necessary to induce germination; other growth regulators such as auxins and cytokinins did not induce a response. Endogenous abscisic acid content in somatic embryos matured at 9% sucrose (487.8 ng/g FW) was approximately double that found in those matured at 3% sucrose (258.4 ng/g FW). This indicates induced dormancy in embryos under high osmotic stress. Alginate encapsulation of embryos facilitated the artificial induction of dormancy to extend the conservation period without germination. The induction of dormancy strengthened resistance to dehydration after the embryos were desiccated to 15% of their normal water content. Reduced chances of embryo survival during long-term desiccation were distinctly delayed in dormant embryos. These results indicate that the induction of dormancy in embryos is a promising application for synthetic seed production.     

Cryopreservation and plant regeneration via somatic embryogenesis using shoot apical domes of mature Pinus roxburghii sarg, trees

Summary The present investigation reports optimized parameters for somatic embryogenesis and cryopreservation of embryogenic cultures using shoot apical domes from mature trees of Pinus roxburghii Sarg. Embryogenic tissue of P. roxburghii Sarg. was cryopreserved for 24 h, 10 d, and 8 wk using sorbitol and dimethylsulfoxide (DMSO) as cryoprotectants. Results indicate that 0.2M sorbitol and 5% DMSO had the best cryoprotecting effect. The recovered tissue showed luxuriant growth on maintenance medium (II). Partial desiccation of thawed embryogenic tissue for 24 h prior to transfer to maturation medium enhanced the maturation of somatic embryos. Maturation frequency increased from 1.3 to 18.3% after 12 h desiccation treatment, and from 18.3 to 61.8% after 24 h of desiccation. However, non-desiccated embryogenic tissue produced the least number of somatic embryos (1.3%) on the maturation medium with the same abscisic acid and Gellan gum concentration. All the three embryogenic lines produced plantlets and had the same appearance and normal growth as compared to unfrozen controls.     

Large Scale Production and Storability of Encapsulated Somatic Embryos of Mothbean (Vigna aconitifolia Jacq)

Storability and germination of sodium alginate encapsulated somatic embryos of Vigna aconitifolia (Jacq.) cv. BMB-43 were tested on half strength Murashige and Skoog (MS) basal medium fortified with coconut water (10% v/v). The frequency of regeneration from encapsulatd embryos was affected significantly by concentration of sodium alginate and the duration of exposure to calcium chloride. Embryos encapsulated with 2.5 % sodium alginate dissolved in MS basal salts solution recorded significantly higher germination than other treatments. A relatively short (5 min) incubation with calcium chloride solution provided uniform encapsulation of embryos that gave the highest percentage (65%) of germination. Synthetic seeds could be stored at 4üC for 50 days without reduction in viability as opposed to non - encapsulated somatic embryos which showed 6% viability after 20 days at 4°C. Germinated synthetic seeds produced normal plantlets.     

Successful long-term preservation of abscisic-acid-treated and desiccated carrot somatic embryos

We have previously reported that strong desiccation tolerance in carrot somatic embryos can be achieved by treatment with abscisic acid (ABA). In this study, we examined the possibility of long-term preservation of ABA-treated and desiccated somatic embryos. Somatic embryos that had been desiccated after treatment with ABA survived for at least 169 weeks at –25 °C. By contrast, somatic embryos that had not been desiccated after treatment with ABA survived for at least 24 weeks at +5 °C but died at –25 °C. Received: 11 July 1998 / Revision received: 20 October 1998 / Accepted: 20 November 1998     

Secondary Somatic Embryogenesis in Abies numidica

The induction of secondary somatic embryogenesis in Abies numidica De Lann. was achieved. Precotyledonary, cotyledonary, and desiccated cotyledonary embryos were used as explants. Cotyledonary embryos before desiccation were the most suitable. The most beneficial was induction medium Schenk and Hildebrandt (SH) with 1 mg dm⁻³ thidiazuron and 1000 mg dm⁻³ myo-inositol. Initiation frequency was from 1 to 34 %. Maturation of somatic embryos was achieved on modified Murashige and Skoog medium supplemented with 40 g dm⁻³ maltose, 100 g dm⁻³ polyethylene glycol-4000 and 10 mg dm⁻³ abscisic acid. Mature somatic embryos after three weeks of desiccation germinated on SH medium with 10 g dm⁻³ charcoal and 10 g dm⁻³ sucrose. This revised version was published online in July 2006 with corrections to the Cover Date.     

白杄体细胞胚胎发生及其植株再生条件的优化研究

为建立白木千体细胞胚胎发生及其植株再生的高频率实验系统 ,对聚乙二醇及干化处理的影响进行了系统研究。结果表明 ,在分化培养基中附加 5 0 g/L聚乙二醇可显著提高愈伤组织的分化频率和每块愈伤组织产生的体细胞胚个数 ,而干化处理又能使体细胞胚的萌发率大幅度上升