非洲紫罗兰的组培快繁技术研究

通过正交试验和单因素试验对非洲紫罗兰的叶片组培快繁技术进行了探讨。结果表明：初代培养基为MS+6-BA1.0mg/L(单位下同)+IBA0.5;诱导丛生芽的适宜培养基为MS+6-BA1.0 +KT0.1 +IBA0.1;有效苗诱导培养基为MS+6-BA0.5+IBA0.5+活性炭0.3%。试管苗生根采用瓶外扦插生根效果较好。     

Regarding the novelty and breeding value of protoplast-derived variants of Russet Burbank (Solanum tuberosum L.)

Summary Striking similarities appear to exist between selected protoplast-derived variants (PD-clones) of the potato cultivar Russet Burbank, and commonly occurring somatic potato sports, called bolters. A limited survey of individuals involved in the certification of Russet Burbank, revealed that bolters do commonly occur in this variety, are highly variable, and sometimes occur at high frequency. It is therefore suggested that bolters and the more promising selected PD-clones of Russest Burbank may be one and the same, arising from the same type of macromutation.One selected PD-clone, three different Russet Burbank bolters, and two controls were analyzed for 15 isozymes in the hope of determining if bolters and promising PD-clones belong to the same class of variants. No isozyme variation between Russet Burbank and its subclones was observed. The breeding value of protoplast-derived variation is discussed in this light.Approved by the Director of the New York State Agricultural Experiment Station for publication as Journal paper No. 3554, Oct. 19, 1983.     

Chromosome numbers of Saintpaulia ionantha plantlets regenerated from leaves cultured in vitro with caffeine and colchicine

Summary Parts of Saintpaulia ionantha leaves were cultured on a medium supplemented with different concentrations of colchicine or caffeine. The explants were transferred to a control medium without these chemicals after different periods of time. The chromosome number of the regenerated plantlets was counted. In all the experiments, polyploid plantlets were found, but the percentage of them was higher with colchicine. There were no differences in frequency of polyploid plantlets among treatments with the same chemical. In both cases a high percentage of cytochimeric plantlets was observed and possible explanations of their occurrence are given.     

A comparison of somaclonal variation in rice plants derived and not derived from protoplasts

To clarify the effect of the type of in vitro culture on the generation of somaclonal variation, protoplast-derived rice plants were compared with rice plants derived from suspension culture or primary calli (not derived from protoplasts). Regenerated plants showing polyploid-like phenotypes appeared at a higher frequency (33–70%) in plants derived from protoplasts than in those not derived from protoplasts (3–6%). In the first progeny (R₁) generation of all regenerated plants, 120 of 368 lines (33%) segregated plants with mutated characters such as albino, dwarf and sterile. In quantitative traits, 62 (21%) and 144 (50%) of 290 Rj lines showed shorter culm and lower seed-fertility, respectively, compared with the control line derived from the selfed seeds of the original cultivar. The frequency of the mutant-possessing R₁ lines among lines derived from protoplasts was not significantly different from those not derived from protoplasts. These results indicate that isolation and culture of protoplasts does not enhance genetic changes other than cytogenetical changes.     

Callus Culture-Derived Somaclonal Variation in the Tropical Pasture Legume Stylosanthes guianensis (Aubl.) Sw.

Plants were regenerated from leaf-derived callus cultures of five genotypes of Stylosanthes guianensis. There were substantial genotypic differences in morphogenetic potential, ranging from 95–100 % of cv.‘Graham' calh to less than 1% of cv,‘Oxley' calh forming shoots. A total of 1024 regenerants (SC₁) were planted out, with 28.1 % surviving to set seed. Variant characters including albinism, reduced fertility, dwarfism and leaf morphology were observed. Variation in selfed progeny (SQ₂) of 138 SC₁ plants was assessed in a replicated field trial involving 2484 SO₂ individuals. Variant phenotypes were observed in 55 (39.9 %) SC₂ families. Many families were segregating for albinism, altered morphology or reduced fertility. Thirty families had lower yields than their parent and earlier and later flowering variants were found. Eight tetraploid families were identified. These were characterized by more erect habit, larger leaves, stems, flowers and seeds, reduced fertility and yield and a tendency to flower later.     

RAPD-based detection of genomic instability in soybean plants derived from somatic embryogenesis

Random amplified polymorphic DNA (RAPD) markers were used to evaluate genetic stability of regenerants of soybean plants obtained through somatic embryogenesis using 180 μM 2,4‐dichloro‐phenoxy acetic acid. Twenty primers were used to screen 44 regenerants from the cultivar ‘Spring’ and 28 from the cultivar ‘CAC‐1’. Three primers were polymorphic for two of the ‘Spring’‐derived regenerants, with a somaclonal frequency of 4.5%. Four primers were polymorphic for the ‘CAC’‐l‐derived regenerant, with a somaclonal frequency of 3.57%. The results indicate the usefulness of RAPD markers to detect genetic instability in soybean primary regenerant plants derived from somatic embryogenesis, and as a certification tool for monitoring genetic stability during the regeneration process.     

Genetic variability in plants regenerated from in vitro culture of sunflower (Helianthus annum L.)

In vitro regenerated sunflower (Helianthus annuus L.) plants (R₁) were self-fertilized and the R₂ generation was evaluated for qualitative traits. A broad range of phenotypic variation was observed and mutation frequencies were calculated. Some in vitro induced variant phenotypes were similar to known spontaneous or induced mutations in sunflower, while others were new. Chlorophyll and carotenoid deficiencies, chimaerical variegation, fasciated stem and capitulum, abnormal shoot development, and other morphological variations, were noted. Substitution of anthers with petaloid structures in a disk-floret mutant indicates a possible homeotic mutation induced by in vitro tissue culture.     

Variation in phenotype and proteins in plants regenerated from cell suspensions of potato cv. BP1

Summary Phenotypic variation, SDS-PAGE and protein-DNA binding were used to determine variation during the in vitro phase of potato plantlets derived from callus and cell suspensions. Of the 27 plantlets assessed. 3 displayed a low or abnormal growth, 16 normal growth which correlated well with the original explant and 9 showed strong or vigorous growth. Differences were not observed in the polypeptide profiles of these plantlets. However distinct differences in the protein-DNA-binding profiles occurred which correlated well with the phenotypic variation observed.     

In vitro Culture of Cucumis sativus

The variability of morphological traits in 1800 individuals of R₂ generation of Cucumis sativus was studied. R₂ generation was obtained from 60 R₁ planes which regenerated from leaf explants of a highly inbred Line of cv. Borszczagowski. All the plants of R₁ population were phenotypically identical with the initial variety; some (8,3 %) of the R₂ progeny, however, exhibited segregations. Variability was manifested in the following traits: 1. type of growth (dwarf and bushy), 2, male sterility, 3, mosaic type leaves, 4, xantha type chlorophyll change. In R₃ generation, only one new phenotype (yellow fruit flesh) has been found. The traits mentioned above were frequently accompanied by lower viability and fertility of R₂ plants. The latter was connected with disturbances in microsporogenesis and lover pollen germination ability. Genic instability of one trait is described.     

Random amplified polymorphic DNA (RAPDs) markers for genetic analysis in micropropagated plants of Robinia pseudoacacia L.

Four years' old micropropagated plants regenerated by enhanced axillary branching from shoot buds of a single genotype of Robinia pseudoacacia were characterized by RAPDs. Random amplified polymorphic DNA analysis was carried outusing 19 random 10-mer DNA primers and 286 RAPD bands were examined which showed 30% polymorphism. Similarity indices ranged from 0.86 to 0.96 among different plants based on RAPD data. The UPGMA dendrogram was constructed based on similarity indices which showed clustering of different plants into subgroups based on similarity values. Our results suggest that somaclonal DNA sequence variations are present even when organized cultures such as shoot buds were used as explant for micro-propagation. This revised version was published online in August 2006 with corrections to the Cover Date.     

Somatic Embryogenesis in Maize and Comparison of Genetic Variability Induced by Gamma Radiation and Tissue Culture Techniques

Sixteen inbred lines and one hybrid of manse were tested for their capability of somatic embryogenesis, and fully developed plants could be regenerated, from ten inbred, lines. The highest frequency of plant regeneration was expressed in the inbred line CHI 31, and when this line was crossed with a recalcitrant, non-regenerating line, the F₁ and BC hybrids were regenerable. The results of reciprocal crosses demonstrated that dominant nuclear genes and cytoplasmic factors are primarily responsible for the heritable determination of embryogenic callus proliferation and in vitro regeneration of maize plants. Somaclonal and radiation-induced variability was studied in maize to assess their nature and potential contribution to plant breeding., The inbred line CHI 31 possessing a high in vitro capacity of somatic embryo formation was used as experiments.] material. CHI 31 plants were selfed and twelve-day old zygotic embryos irradiated with ⁶⁰Co gamma radiation in situ. Mature caryopses were harvested and assigned as M₁ material. In another series, immature zygotic embryos (size 1.2—1.5 mm) were cultured in vitro on N-6 medium supplemented with 2,4-D (2.5 μM), and somatic embryos regenerated into plants; these were transplanted into soil and self-pollinated. Regenerants from non-irradiated cultures were grown as R₁ generation, while regenerants from irradiated explants were considered as M₁R₁ generation. The genetic variability was evaluated in the M₂, R₂ and M₂R₂ generations, respectively, and compared with a non-treated seed control. Irradiation induced a variety of chlorophyll and morphological variants segregating in the M; generation; however, the frequency of deviant types obtained in the R: generation (somaclonal variation) was significantly exceeding the one derived from the M₂ populations. The combination of expert irradiation and in vitro regeneration was most effective for the manifestation of chlorophyll and morphological o if types in the M₂R₂ generation, and increased drastically the frequency of early flowering variants. Differences in the segregation patterns of mutant phenotypes amonsister somaclones in the R₃ and M₃R₃ generations indicate a different genetic basis, of plants originating from the same explant. This phenomenon suggests a mutational sectoring of the callus during culture. Radiation induced and somaclonal variation exerted a similar spectrum of chlorophyll and morphological deviants.     

Plant regeneration from protoplasts of Iris hollandica Hort.

The effects of glucose concentrations, different sugars and combinations of 2,4-D and kinetin on cell division and colony formation were examined in cultures of protoplasts isolated enzymatically from suspension cultures of Iris hollandica N6 medium supplemented with 1 mg/l 2,4-D, 1 mg/l kinetin, 200 mg/l casein hydrolysate, 250 mg/l proline, 0.3– 0.5 M glucose and 20 g/l agarose was suitable for cell division and colony formation. When colonies formed were transferred to hormone-free MS medium, many shoots were induced. In addition, when induced shoots were transferred to MS medium with 1 mg/l NAA, root induction was observed. A plant regeneration system from protoplasts of I. hollandica was thus established.     

Effect of Mutagenic Treatments on Somaclonal Variation in Wheat (Triticum aestivum L.)

Different wheat genotypes were treated with gamma-rays, sodium azide (SA) and EMS before tissue culture and immature embryos from M₁ plants or plants shortly after exposure to gamma-rays were used to initiate callus culture. Thousands of plants were regenerated and used to investigate the effect of mutagenic treatments on the regenerated plants and somaclonal variation in the M₃R₂ and M₂R₂ generations. The results showed that mutagen-induced damage in terms of reduction in plant height, fertility and spike length were not outstanding in the regenerated plants as compared with the untreated control. In the M₃R₂ generation, only SA treatment had significantly higher frequencies of somaclonal variations than the control. Increases in the variation frequencies were observed when explant embryos were irradiated with 2.5 and 5 gy gamma-rays and the highest frequency appeared when embryos were exposed to 5 gy gamma-rays on the 5th day after anthesis. Increased variation spectra also resulted from mutagenic treatments and most of the variants recovered were unsuitable for plant improvement.     

Somaclonal variation in cell suspension culture-derived regenerants of Cymbopogon martinii (Roxb.) Wats var. motia

Plants regenerated from cell suspension cultures of palmarosa grass, Cymbopogon martinii (Roxb.) Wats, were analysed for somaclonal variation in five clonal generations. A wide range of variation in important quantitative traits, e.g. plant yield, height, tiller number, oil content and qualitative changes in essential oil constituents—geraniol, geranyl acetate, geranyl formate and linalool—were observed among the 120 somaclones screened. Eight somaclones were selected on the basis of high herb and oil yield over the donor line and high geraniol content in the oil. Based on performance in the field trials, three superior lines were selected, and maintained for five clonal generations. The superior lines exhibited a reasonable degree of stability in the traits selected.     

Field performance of embryogenic cell suspension-derived banana plants (Musa AAA, cv. Grande naine)

Growth and yield characteristics of two different clones of banana plants (Musa AAA cv. Grande naine) originating from four months old embryogenic cell suspensions were studied. These characteristics were compared with those plants produced by the conventional in vitro budding multiplication method. Two types of variants were observed during the acclimatization phase among 500 embryogenic cell suspension derived plants. The first type related to banana plants with `variegated or deformed leaves' were also observed in in vitro budding derived plants. The second type concerned `fasciated-leafed' plants. During the field growth, these two variant types produced plants morphologically similar to the other plants. Thus, none of the cell suspension derived plants exhibited off-type traits in the field. A Fisher block model was used to compare the field performances of the two clones produced through the two in vitro propagation techniques. The analysis of variance showed that there were no significant differences between the plants produced by either micropropagation techniques for the plant height and circumference, the length of the reference leaf, the number of nodal clusters of the inflorescence and of fruits, the bunch weight, the period of time between planting and flowering, and between planting and harvesting. This study showed that banana plants with an agronomical behaviour similar to those produced by the conventional in vitro budding method could be regenerated from embryogenic cell suspension. This revised version was published online in July 2006 with corrections to the Cover Date.     

Fertility and growth in the field of Lolium perenne and Festuca rubra plants regenerated from suspension cultured cells and protoplasts

The effect of regeneration of Lolium perenne and Festuca rubra from embryogenic suspension cells and protoplasts on fertility and growth was evaluated. Embryogenic suspension cultures were either routinely subcultured or cryopreserved and re-established. Phenology, morphology and fertility of regenerated plants were studied for two growing seasons in a replicated field experiment. Most regenerated L. perenne and F. rubra plants showed a delay in inflorescence emergence, a reduced seed yield and differences in morphological traits when compared with seed-grown plants. For L. perenne, performance of plants regenerated from cryopreserved suspension cultures and protoplasts was similar to that of respective plants regenerated from routinely maintained suspension cultures. However, differences in performance were observed for respective regenerants in F. rubra. The phenotypic deviation observed was partly reflected in the randomly amplified polymorphic DNA (RAPD) analysis performed. However, regenerants of both species showing similar, or even superior performance to the seed-grown plants were also found. Embryogenic suspension cells and corresponding protoplasts of L. perenne and F. rubra have the potential for producing fertile, well-performing plants which can be integrated in breeding programs.     

Modification of oilseed rape to produce oils for industrial use by means of applied tissue culture methodology

Summary A programme of research was designed to investigate methods for the modification of the fatty acid profiles of high performance lines of oilseed rape (Brassica napus L.) in an attempt to produce lines with enhanced levels of industrially useful fatty acids. The methodology employed to achieve these objectives was based on the exploitation of somaclonal or protoclonal variation, and targeted somatic hybridization using wild cruciferous germplasm as fusion partners.A range of somaclonal lines was produced from shoot regeneration protocols. These lines underwent replicated, randomised glasshouse trials for morphological assessment followed by gas chromatographic analysis to monitor any changes in fatty acid profile. It was found that a small number of lines exhibited potentially useful changes in oleic acid and polyunsaturated fatty acid content. Protoplast regeneration and electrofusion protocols for a range of winter oilseed rape lines were developed, and methods for the isolation and fusion of protoplasts of the wild crucifer Lunaria annua (chosen for its high nervonic acid content) established.Abbreviations GC Gas Chromatography     

Somaclonal Variation in Tissue Culture of Triticum aestivum×Agropyron desertorum F1 Hybrid

Callus induced from immature inflorescences of the partially self-fertile hybrids (2 n = 35; ABDPP) between Triticum aestivum (2n = 42; AABBDD) and Agropyron desertorum (2n = 28; PPPP) led to the regeneration of 88 plants on MS medium supplemented with 2 mg/l of 2,4-D. These regenerants were used to investigate somaclonal variation and to obtain more selfed derivatives. Immature inflorescences at the stage of developing floral primordia gave the best response in terms of callus induction and plant regeneration. The regenerants exhibited great variability for most morphological traits. Although the regenerants did not exhibit variation in chromosome number, they did show a higher degree of meiotic instability than the initial hybrid. In particular, the regenerants gave much higher selfed seed-set (5.49 %) than the donor hybrid (0.46 %), so that a total of 484 selfed seeds were obtained.     

Frost Resistance of Somaclones Derived from Triticum aestivum L. Winter Wheat Calli

Frost resistance was studied in SC₄ seedlings generated by self pollination from 31 (SC₄) plants of ‘GK Csongor’ winter wheat variety derived from resistance than ‘GK Csongor’. With respect to percentage survival, one family possessed significantly higher frost resistance as compared to the control at a temperature of -13°C In the case of regrowth analysis, 22 of the 31 families showed less growing capacity and 5 proved to be significantly better than ‘GK Csongor’. According to both testing methods, one family showed significantly higher frost resistance than the control.     

木豆高频不定芽再生及体细胞遗传变异的分子检测

以木豆子叶节为外植体,建立高频不定芽再生体系,并对继代丛芽的遗传变异进行分子检测。结果表明:子叶节不定芽再生的最佳培养基为EC6附加4.0mg/LBA和0.1mg/LIBA;丛芽增殖培养以EC6附加4.5mg/LBA、0.1mg/LIBA和0.4mg/LKT效果最佳。同时,随机挑选继代第5代的20个丛芽提取基因组DNA,采用45个ISSR引物对基因组DNA进行分子检测,结果显示,在所用引物范围内均未检测到变异。