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Progress made in FEC transformation of cassava 总被引:2,自引:0,他引:2
Krit Raemakers Marianne Schreuder Isolde Pereira Tichafa Munyikwa Evert Jacobsen Richard Visser 《Euphytica》2001,120(1):15-24
In cassava friable embryogenic callus (FEC) has been used to obtain transgenic plants using particle bombardment, electroporation,
and Agrobacterium tumefaciens. FEC cultures have been obtained in 6 of the10 tested genotypes. In all genotypes FEC could be regenerated into plants,however
the efficiency differed between the genotypes. Almost all plants regenerated from 6 months old FEC cultures of TMS604444,
Adira 4,Thai 5 and M7 were morphological similar to control plants. However, in R60 and R90 a large number of plants were
not identical to control plants. Older FEC lines of TMS60444 have a reduced ability to regenerate plants and the plants show
somaclonal variation. Somaclonal variation is observed in the same extend in transgenic and non-transgenic plants. The origin
of this variation is both genetic and epigenetic. Luciferase based selection is less efficient in producing transgenic lines
than chemical selection. Furthermore Agrobacterium tumefaciens mediated transformation is much more efficient than particle bombardment with respect to the production of transgenic lines.
A tentative model is introduced which best describes the effect of different selection regimes on the time period required
to produce transgenic plants. Kanamycin and stringent luciferase selection required a shorter period of time than selection
based on hygromycin, phosphinothricin or non-stringent luciferase. However, a more significant reduction of time was obtained
if young instead of old FEC lines of genotype TMS60444 were used for genetic modification. In accordance to the model these
young FEC lines of TMS60444 produced transgenic plants within 4 months with both Agrobacterium tumefaciens combined with kanamycin selection and particle bombardment combined with stringent luciferase selection.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
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Efficient production of transgenic plants by Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) 总被引:1,自引:0,他引:1
An efficient and reproducible method was developed for Agrobacterium-mediated transformation of embryogenic suspension cultures of cassava. LBA4404(pTOK233), containing the nptII, hph and gus marker genes, was used in the experiments. Chemical selection by means of kanamycin was used to establish 1037antibiotic
resistant callus lines, of which 526 showed GUS expression. Of the 241 callus lines that were transferred to maturation medium
219formed somatic embryos. Thirty-seven of the 38 lines that were transferred to germination medium produced plants. GUS-positive
plants could be obtained from 31 lines; in 14 of those lines 100% of the produced plants were GUS-positive, the remaining
17 lines yielded GUS-positive plants at an average of 72%. The transgenic nature of these plants was confirmed by Southern
blot analysis.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
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Regeneration and transformation of cassava 总被引:3,自引:0,他引:3
A prerequisite for the development of a successful transformation system is the availability of efficient regeneration systems.
Up to 1995 the only available regeneration system in cassava was an organized type of somatic embryogenesis. Transformation
of these organized somatic embryogenic cultures with particle bombardment or Agrobacterium tumefaciens resulted in chimeric
transformed embryos. However, the transformed sector was lost after repeated cycles of secondary somatic embryogenesis. After
1995 a less organized system of somatic embryogenesis was developed, so called friable embryogenic callus (FEC) and a system
of adventitious shoot regeneration. The FEC regeneration system was combined successfully with particle bombardment. Selection
of transgenic plants was based on either luciferase activity, or resistance to the aminoglycoside paromomycin or the herbicide
phosphinothricin. Furthermore, protoplasts of FEC are able to regenerate into plants and can be transformed by electroporation.
The adventitious shoot regeneration system was combined successfully with Agrobacterium tumefaciens. For this mature somatic
embryos were cocultivated with Agrobacterium and cultured for adventitious shoot development. After selection based on the
aminoglycoside geneticin or on hygromycin transgenic plants were formed.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
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C.E. van Schaik C. van der Toorn M.J. De Jeu C.J.J.M. Raemakers R.G.F. Visser 《Euphytica》2000,115(1):17-26
The successful application of plant biotechnology to Alstroemeria improvement will largely depend on the availability of an efficient regeneration/transformation system. Regeneration in Alstroemeria is accomplished from nodular embryogenic callus initiated from zygotic embryos. Histological studies of embryogenic callus
initiation from 4-weeks old cultured ovules revealed that the outermost layers of the protoderm of the embryogenic nodules
divided to form either a new nodule or aproembryo. Transient gene expression after particle bombardment of nodular embryogenic
callus was optimized using DNA of pAHC25. The highest β-glucuronidase expression was found when the GUS gene was under control of the maize ubiquitin promoter, the target tissue was placed 5 cm below the microcarrier launch assembly
and when the rupture disc-breakage point was between 650–900 psi. Kanamycin blocked regeneration of somatic embryos, however,
did not block growth of nodular embryogenic callus. With phosphinothricin both callus growth and regeneration were blocked.
Bombardment of nodular embryogenic callus with DNA of pAHC25 combined with selection on medium containing phosphinothricin
resulted in putative transgenic chimeric. Friable calli were selected from nodular embryogenic callus and used to initiate
suspensions. These cell suspensions were subjected to transformation by particle bombardment using DNA of pAHC25 and resulted
in a stable transformed friable callus line after selection based on luciferase activity. Even after 2 years of maintenance
this callus line was luciferase positive and the Polymerase Chain Reaction analysis demonstrated the presence of the introduced
gene in this friable callus line.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
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Secondary somatic embryogenesis and applications in plant breeding 总被引:17,自引:0,他引:17
Summary Secondary somatic embryogenesis is the phenomenon whereby new somatic embryos are initiated from somatic embryos. Such cultures have been described in at least 80 Gymnosperm and Angiosperm species. In the initial step (primary somatic embryogenesis) such cultures have to be started from plant explants. In general, primary somatic embryogenesis from vegetative plant explants is, indirect and mostly driven by auxin (AUX) or auxin and cytokinin (AUX/CYT) supplemented media, whereas, from zygotic embryos it is direct and driven, to a larger extent, by CYT or growth regulator free media. Primary somatic embryogenesis from floral plant explants is between these two extremes. Indirect and direct somatic embryogenesis should be seen as two extremes of one continuum: in indirect somatic embryogenesis the embryos develop up to the (pre)-globular stage and in direct somatic embryogenesis to mature stages before they are subjected to secondary embryogenesis. In general, secondary embryogenesis requires no growth regulators in species with CYT driven primary embryogenesis. Whereas, continuous exposure to growth regulators is needed in species with CYT/AUX or AUX driven primary embryogenesis.In most species somatic embryos can be converted into shoots, although the frequencies are mostly low. In general, somatic embryos induced by growth regulator free or CYT supplemented media meet more difficulties in shoot development than embryos induced by AUX supplemented media. Applications of secondary somatic embryogenesis for plant breeding are discussed. 相似文献
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