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1.
Fertile plants have been obtained from maize (Zea mays L.) embryogenic suspension culture protoplasts. Friable, embryogenic callus initiated from an immature embryo from a cross involving the genotypes A188, B73, and Black Mexican sweetcorn was used to establish a rapidly growing embryogenic suspension culture. After nine months in culture, high yields of viable protoplasts (30×106/ gram fresh weight) were obtained following a 1.5 hour enzymatic digestion. Protoplasts cultured with feeder cells divided and formed embryogenic callus, from which male and female fertile plants were regenerated. Protoplast-derived R1 plants were self-pollinated and immature R2 embryos isolated for callus initiation. Female fertile plants have also been produced from protoplasts isolated from an R2-derived suspension culture. Significant interactions between protoplast and feeder-cell lines were observed.Abbreviations BC
backcross
- BMS
Black Mexican Sweetcorn
- 2,4-D
2,4-dichlorophenoxyacetic acid
- PWS
protoplast wash solution (0.2 M mannitol, 80 mM CaCl2)
- FDA
fluorescein diacetate
- ABA
abscisic acid 相似文献
2.
Summary Plants were regenerated from maize (Zea mays L.) protoplasts isolated from embryogenic cell suspensions. The donor maize suspension cultures were established from friable callus initiated from microspores of a commercial supersweet hybrid (sh2sh2). The frequency of cell colony formation was higher when protoplasts were cultured on feeder layers of maize cells as compared with a liquid thin layer method. It was demonstrated that haploid and dihaploid soil-grown plants can be regenerated from maize protoplasts isolated from haploid cell cultures. 相似文献
3.
Transient gene expression after electroporation of protoplasts derived from embryogenic maize callus
Protoplasts isolated from embryogenic callus cultures derived from immature embryos ofZea mays L. are suitable for analysis of transient gene expression using electroporation-mediated DNA transfer. Expression of introduced genes is comparable to the levels obtained with protoplasts from Black Mexican Sweet suspension cultures. Two different promoters, that directing synthesis of the 35S RNA of cauliflower mosaic virus and the maizeAdh1 promoter were placed in front of the luciferase reporter gene to assess protoplast gene expression and the impact of an intron on expression level.Abbreviations 35S
promoter isolated from CaMV
- CaMV
cauliflower mosaic virus
-
Adh1
maize gene encoding Alcohol dehydrogenase-1 enzyme
- BMS
suspension cultures of the Black Mexican Sweet maize variety 相似文献
4.
Wang Xiao Xue-Lin Huang Xia Huang Ya-Ping Chen Xue-Mei Dai Jie-Tang Zhao 《Plant Cell, Tissue and Organ Culture》2007,90(2):191-200
A protocol for plant regeneration from protoplasts of Musa acuminata cv. Mas (AA) via somatic embryogenesis was developed. Viable protoplasts were isolated from embryogenic cell suspensions
at a yield of 1.2 × 107 protoplasts/ml packed cell volume (PCV). Liquid and feeder layer culture systems with medium-A and medium-B were used for
protoplast culture. In liquid culture system, medium-B was more efficient for inducing cell division (17.5% at 14 days) and
colony formation (6.7% at 28 days) than medium-A. However, all protoplast-derived cell colonies (PDCC) obtained from liquid
culture system could not develop further. In feeder layer culture system, there was no significant difference between medium-A
and medium-B on cell division and colony formation of the cultured protoplasts, and the cell division frequency at 14 days
and colony formation frequency at 28 days were 24.5% and 11.2%, respectively, in medium-B. Comparative study on the effects
of BAP (2.2 μM, 4.4 μM, 8.8 μM), zeatin (0.4 μM, 0.8 μM, 1.2 μM) and TDZ (0.2 μM, 0.4 μM, 0.6 μM) on embryo formation of PDCC
from feeder-layer culture indicated that TDZ was best. TDZ at 0.4 μM induced 7906 mature embryos per ml PCV PDCC, which was
4-fold the frequency as with BAP at 4.4 μM, 7.5-fold as with zeatin at 0.8 μM and 150-fold as control medium (no mentioned
cytokinins) after 45 days on M3 medium. About 44% of the mature embryos were converted into plantlets with poor root system
after subculture on M4 medium. Root further development of regenerated plantlets was promoted by addition of activated charcoal
(AC) to MS basal medium. 相似文献
5.
Development and Characterization of a Nonmorphogenetic Cell Line of Wheat (Triticum aestivum) 总被引:1,自引:1,他引:0
Arron C. Guenzi Kay Scheets J. Larry Green John P. Fellers 《Plant Cell, Tissue and Organ Culture》2004,78(1):23-28
This study was conducted to compare characteristics of a wheat (Triticum aestivum L.) cell line to those of the maize (Zea mays L.) black Mexican sweet (BMS) cell line and to compare protoplasts isolated from suspension cells of these cell lines. The
wheat cell line was established from immature-embryo derived callus of the experimental line ‘ND7532’ and was conditioned
for growth in suspension culture. For both cell lines, measurements of packed cell volume (PCV), fresh weight (FW), and dry
weight (DW) were taken at 3 day intervals from suspension cultures. Measurements of FW of calluses cultured from suspension
cells of both cell lines were taken at 6 day intervals. The morphogenetic potential of the wheat ND7532 cell line was tested
in both callus and suspension cultures using media promoting regeneration and/or organogenesis. Growth rates of ND7532 cells
in suspension culture were comparable to those of BMS cells. However, relative growth rates of calluses recovered from ND7532
suspension cells were slower than those of calluses recovered from BMS suspension cells. The ND7532 cell line has very limited
morphogenetic potential and has been maintained as rapidly growing callus tissue for 11 years. Yields of protoplasts from
suspension cells of the two cell lines were comparable, though ND7532 protoplasts were typically smaller. The wheat cell line
has is now designated ND7532-NM (nonmorphogenetic) and is available for cellular and molecular biology research. 相似文献
6.
Protoplasts were released from embryogenic suspension culture of Paspalum scrobiculatum and cultured in either liquid or semisolid KM medium supplemented with 2,4-D in the dark at 24°C with or without a feeder layer. Cell wall formation was observed in 75% of the plated protoplasts. Microcolonies developed after 10 d of culture, which in turn formed callus upon transfer to M-2 medium (Nayak and Sen, 1989). The highest plating effeciency (ca 7%) was obtained in thin-layer liquid culture. The macrocalli formed somatic embryos which regenerated to plantlets. The plantlets were grown to flowering plants upon transfer to soil.Abbreviations BAP
6-benzylaminopurine
- 2,4-D
2,4-dichlorophenoxyacetic acid
- FDA
Fluoresceine diacetate
- MES
2-(N-Morpholino) ethanesulfonic acid
- MS
Murashige & Skoog medium (1962) 相似文献
7.
Sugar beet protoplasts (Beta vulgaris L.) were isolated from hypocotyl-derived suspension cells and cultured on modified Murashige and Skoog medium supplemented
with 5 μM naphthaleneacetic acid (NAA) and 2 μM 6-benzyl-aminopurine (BAP). Protoplasts were plated at a density 1.0–1.5×105 cm−3 and incubated in either liquid medium or in medium solidified by 1.2% agarose, at 25°C in the dark. Comparison of two methods
of culture unequivocally showed the second to be superior. Immobilizing the protoplast in agarose proved to be essential for
obtaining sustained protoplast division and reproducible colony formation. The plating efficiency after two weeks of culture,
expressed as the percentage of protoplasts which developed to form colonies, reached 40%. Subsequent subcultures of protoplast-derived
callus to regeneration media with different concentrations of BAP (5 μM, 10 μM, 20 μM, 30 μM) resulted in very good callus
proliferation at the three lowest concentrations, although organogenesis was not achieved. 相似文献
8.
Paul W. J. Taylor Hian-Lien Ko Stephen W. Adkins Carl Rathus Robert G. Birch 《Plant Cell, Tissue and Organ Culture》1992,28(1):69-78
For 18 sugarcane cultivars, four distinct callus types developed on leaf explant tissue cultured on modified MS medium, but only Type 3 (embryogenic) and Type 4 (organogenic) were capable of plant regeneration. Cell suspension cultures were initiated from embryogenic callus incubated in a liquid medium. In stage one the callus adapted to the liquid medium. In stage two a heterogeneous cell suspension culture formed in 14 cultivars after five to eight weeks of culture. In stage three a homogeneous cell suspension culture was developed in six cultivars after 10 to 14 weeks by selective subculturing to increase the proportion of actively dividing cells from the heterogeneous cell suspension culture. Plants were regenerated from cell aggregates in heterogeneous cell suspension cultures for up to 148 days of culture but plants could not be regenerated from homogeneous cell suspension cultures. High yields of protoplasts were obtained from homogeneous cell suspension cultures (3.4 to 5.2 × 106 protoplasts per gram fresh weight of cells [gfwt-1]) compared to heterogeneous cell suspension cultures (0.1 × 106 protoplasts gfwt-1). Higher yields of protoplasts were obtained from homogeneous cell suspension cultures for cultivars Q63 and Q96 after regenerating callus from the cell suspension cultures, then recycling this callus to liquid medium (S-cell suspension cultures). This process increased protoplast yield to 9.4 × 106 protoplasts gfwt-1. Protoplasts isolated from S-cell suspension cultures were regenerated to callus and recycled to produce SP-cell suspension cultures yielding 6.4 to 13.2 × 106 protoplasts gfwt-1. This recycling of callus to produce S-cell suspension cultures allowed protoplasts to be isolated for the first time from cell lines of cultivars Q110 and Q138. 相似文献
9.
Conditions have been developed that induce maize (Zea mays L.) protoplasts to re-synthesize cell walls and to initiate cell divisions. Two types of embryogenic maize callus were used as a source of protoplasts: a heterogeneous callus (Type I) derived from immature embryos after three weeks in culture, and a friable, rapidly growing callus (Type II) selected from portions of the Type I callus. Many variables in the growth conditions of the donor tissue (type of medium, transfer schedule, age of callus), protoplast isolation solutions (pH, osmolarity, type and concentration of cell wall hydrolyzing enzymes, addition of polyamines) and conditions (amount of time in enzyme, amount of tissue per volume of enzyme incubation medium, agitation, preplasmolysis of source tissue, type of callus), and purification procedures (filtration and-or flotation), were found to affect both yield and viability of protoplasts (based upon fluorescein-diacetate staining). Our isolation procedure yielded high numbers of viable, uninucleated maize callus protoplasts which were densely cytoplasmic and varied in size from 20 to 50 m in diameter. Protoplasts plated in solid medium formed walls and divided several times. Of several gelling agents tested for protoplast propagation, only agarose resulted in protoplasts capable of sustained divisions leading to the formation of microcalli. Plating efficiency was established over a wide range of protoplast densities (103–107 protoplasts/ml). Highest plating efficiency (25%) was obtained at 1·106 protoplasts/ml). The resulting microcalli grew to be dense clusters of about 0.1–0.5 mm in diameter and then stopped growing. Nurse cultures of maize and carrot (Daucus carota L.), were used to establish that individual protoplasts (not contaminating cells or cell clusters) formed walls and divided. Nurse cultures also increased the efficiency of microcallus formation from protoplasts.Abbreviations 2,4-D
2,4-dichlorophenoxyacetic acid
- MS
Murashige and Skoog (1962) salts
- MS 1D
Murashige and Skoog salts with 1 mg/l 2,4-D
- MS 2D
Murashige and Skoog salts with 2 mg/l 2,4-D
- N6
medium of Chu et al. (1975)
- NN67-mod
medium of Nitsch and Nitsch (1967) as modified in the present paper
- FDA
fluorescein diacetate
- LMP
low melting point 相似文献
10.
Viable protoplasts of Taxus yunnanensis were isolated from friable, light yellow callus. Protoplast yield was dependent on callus age, with a maximum from 20-day-old
callus. Protoplasts were induced to undergo sustained divisions and to form cell colonies when cultured in medium consisting
of B5 salts, KM vitamin and organic components, 0.45 M fructose, 3.0 mg l-1 2,4-dichlorophenoxyacetic acid and 0.1 mg l-1 kinetin. The planting density was 2.5–3.0×105 protoplasts per ml of culture medium. Cell-free extract from callus enhanced protoplast division and the highest plating
efficiency was about 7%. Protoplast-derived colonies showed significant variations in both growth and paclitaxel content.
A negative correlation existed between paclitaxel accumulation in colonies and their growth to some extent (r = −0.4485).
Among 70 colonies isolated from the heterogeneous protoplast cultures, colony TY-7 accumulated the highest paclitaxel content.
Paclitaxel accumulation in colony TY-7 was not great enough to produce paclitaxel for commercial purposes, however, success
in inducing colony formation from T. yunnanensis protoplasts provides an opportunity to obtain cell lines with high paclitaxel productivity from mutagenized protoplast cultures.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献