Role of genetic background in somatic embryogenesis in Medicago

Brown, Daniel C.; Atanassov, A.

doi:10.1007/bf00042269

Cited by 152 publications

(76 citation statements)

References 16 publications

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“…Leaves of actively growing glasshouse plants were sterilized (5 sec in 70% ethanol followed by 12 min in 1 YO sodium hypochlorite) and washed three times with sterile distilled water. Leaf explants (4 mm2) were submerged in the bacterial suspension for 10 min, then lightly blotted and placed on B5h medium (Brown and Attanosov, 1985) with a 12 h photoperiod of fluorescent light at 26°C for 3 days of co-cultivation. Leaf segments were then washed three times with sterile water and excess water was shaken off before the segments were transferred to selective medium.…”

Section: Plant Transformation and Regenerationmentioning

confidence: 99%

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Vicilin with carboxy‐terminal KDEL is retained in the endoplasmic reticulum and accumulates to high levels in the leaves of transgenic plants

et al. 1992

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SummaryGene constructs were designed to test the effect of the endoplasmic reticulum (El?)-targeting signal, KDEL, on the level of accumulation of a foreign protein in transgenic plants. The gene for the pea seed protein vicilin was modified by the addition of a sequence coding for this tetrapeptide at its carboxyl terminus. The altered gene was placed under the control of a CaMV 35s promoter and its expression in the leaves of both tobacco and lucerne (alfalfa) was compared with that of an equivalent vicilin construct lacking the KDELcoding sequence. The presence of the ER-targeting signal led to a greatly enhanced accumulation of the heterologous protein. In lucerne and tobacco leaves, the level of vicilin-KDEL protein was 20 and 100 times greater than that of the unmodified vicilin, respectively. These differences in expression level could not be explained by corresponding differences in the steadystate levels or the translatability of the mRNAs. However, when the stability of vicilin and vicilin-KDEL proteins was compared in their respective transgenic hosts, unmodified vicilin was found to be degraded with a half-life of 4.5 h while vicilin-KDEL was much more stable with a half-life of more than 48 h. lmmunogold labelling of leaf tissues from transgenic lucerne and tobacco showed the presence of vicilin associated with large aggregates within the ER lumen of vicilin-KDEL plants. No such aggregates were detected in transgenic plants expressing wild-type vicilin.It is concluded that the carboxy-terminal KDEL caused the retention of the modified vicilin in the ER, and that this retention led to the increased stability and higher level of accumulation of vicilin-KDEL in leaves of transgenic plants.

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Section: Plant Transformation and Regenerationmentioning

confidence: 99%

“…Kanamycin-resistant embryos emerged within the next 3 weeks. The embryos were transferred to SHe medium (Brown and Attanosov, 1985) without selection to allow embryo maturation and plantlet formation.…”

Section: Plant Transformation and Regenerationmentioning

confidence: 99%

Vicilin with carboxy‐terminal KDEL is retained in the endoplasmic reticulum and accumulates to high levels in the leaves of transgenic plants

et al. 1992

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“…This clonal line is a highly embryogenic genotype isolated from cv. Rangelander (Brown and Atanassov 1985;Meijer and Brown 1987a,b). Donor plants were propagated in vitro by subculturing individual nodes in 75-mL Schenk and Hildebrandt (1972) medium modified to contain 29.2 mM sucrose, 0.1 mM indole-3-acetic acid, 0.1 mM iso-pentyl adenine, 1% Bactoagar (Difco), pH 5.9, in 10-cm Magenta containers (Magenta Corp., Chicago, IL).…”

Section: Donor Plantsmentioning

confidence: 99%

“…These 10 genotypes were selected from a wide range of genetic backgrounds. Line A70.34 is a commonly used line for somatic embryogenesis in a number of laboratories (Brown and Atanassov 1985;Meijer and Brown 1987a,b), but this genotype was reported to be difficult to transform with Agrobacterium (Du et al 1994). Line 11.9 was screened as a highly responsive genotype for Agrobacterium transformation (Desgagnes et al 1995) At least three replicates (one Petri dish typically containing 8-10 explants or a measured volume of cell sample was considered to be a replicate) were used for the bombardment for each staged tissue or culture to obtain expression measurements.…”

Section: Stage IImentioning

confidence: 99%

Transient expression of a reporter gene changes significantly during somatic embryogenesis in alfalfa

Tian¹,

Brown²,

Webb³

2000

Can. J. Plant Sci.

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Transient expression of a reporter gene changes significantly during somatic embryogenesis in alfalfa. Can. J. Plant Sci. 80: 765-771. Transient expression of the β-glucuronidase (GUS) gene driven by a 35S constitutive promoter was investigated during somatic embryo development in alfalfa (Medicago sativa L.) following microprojectile bombardment. The level of gene expression varied with the different developmental stages and changed dramatically within the stages. During embryogenic callus induction (stage I), expression was low in explants freshly transferred to culture, but increased when cells started to divide and show competence for somatic embryogenesis. Gene expression, however, decreased when extensive callus formation began and reached the lowest observed level when callus was fully developed. Reporter gene expression was consistently low during embryogenic cell proliferation (stage II). In contrast, high levels of reporter gene expression were detected in non-embryogenic suspension cultures during cell proliferation. Extremely low reporter gene expression was detected in embryonal tissues and young embryos at the embryo development and maturation stage (stage III). Expression increased as the somatic embryos developed and reached the highest level when the embryos reached maturity. Transient expression levels were similar in excised petiole tissue across 10 alfalfa genotypes, which included both diploid and tetraploid genotypes. In general, reporter gene expression was found to be higher in more organised mature tissues and organs than in less-organised or young tissues. The developmental status of the cells and tissues appears to be an important factor in the degree of transient expression of the introduced gene. Changes of gene expression during embryogenesis and factors affecting gene expression are discussed. Nous avons examiné l'expression transitoire du gène codant pour la β-glucuronidase (GUS) entraîné par un gène promoteur constituant 35S après bombardement par micro-projectiles durant le développement somato-embryonnaire de la luzerne (Medicago sativa L.). En plus de varier selon le stade de développe-ment, le niveau d'expression du gène changeait de façon spectaculaire à l'intérieur d'un même stade. Pendant l'induction du cal embryogénique (stade I), il était faible dans les explantats fraîchement transférés dans le milieu de culture, mais il augmentait lorsque les cellules commençaient à se diviser et à manifester une compétence d'embryogenèse somatique. En revanche, il diminuait au début de la phase de callogénèse, extensive pour enfin atteindre son plus bas niveau quand le cal arrivait à son plein développement. L'expression du gène rapporteur demeurait faible durant la phase de prolifération des cellules embryogéniques (stade II), alors qu'elle atteignait un degré élevé durant la phase de prolifération des cellules non-embryogéniques. Un très faible degré d'expression du gène rapporteur était observé dans les tissus embryonnaires et dans les jeunes embryons aux stades de développement et...

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“…Brown and Atanassov (1985) evaluated numerous alfalfa cultivars to determine the role of genetic background in the in vitro culture of Medicago. After 28 d on B 5 h medium, a wide range of callus yield was observed among cultivars ranging from 62 mg for cv.…”

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confidence: 99%

Quantitative genetic analysis of in vitro callus proliferation in alfalfa

Kielly¹,

Bowley²

1997

Can. J. Plant Sci.

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Kielly, G. A. and Bowley, S. R. 1997. Quantitative genetic analysis of in vitro callus proliferation in alfalfa. Can. J. Plant Sci. 77: 225-229. Genetic variability exists for in vitro callus growth within plant species, including alfalfa (Medicago sativa L.). The objective of this study was to investigate, using diallel crosses, the genetic variance components of in vitro callus production in tetraploid alfalfa. Nine genotypes were randomly chosen from among 60 plants from the cultivar Saranac and the experimental population OAC81-120R, which were previously characterized as being able to initiate callus. The group of nine plants was divided and crossed to generate two, six-plant diallel sets (A and B) with reciprocals; and the progeny were evaluated for callus production. Significant (P < 0.01) differences were found among crosses and for general combining ability (GCA) effects in both diallels. Specific combining ability (SCA) and reciprocal effects were found to be nonsignificant (P > 0.05). Estimates of heritability (h 2 ) for callus production on a plot mean basis were 73% and 61% for diallel A and B, respectively. Since the heritabilities were high, and no reciprocal effects were detected, breeding strategies employing recurrent selection should result in the development of high in vitro callus producing populations. Key words:Medicago sativa L., callus proliferation, tissue culture, quantitative genetics, heritability, alfalfa Kielly, G. A. et Bowley, S. R. 1997. Analyse génétique quantitative de la callogénèse in vitro chez la luzerne. Can. J. Plant Sci. 77: 225-229. Il existe de la variabilité génétique relativement au développement du cal in vitro chez les espèces végétales, notamment, la luzerne (Medicago sativa L). L'objet des présentes recherches était d'examiner par voie de croisements dialléliques les composantes de la variance génétique relative à la callogénèse in vitro chez la luzerne tétraploïde. Nous avons choisi au hasard neuf générations à partir d'environ 60 plantes du cultivar Saranac et de deux populations expérimentales OAC81-120R précédem-ment caractérisées comme capables de générer des cals. Les neufs plantes étaient séparées puis croisées de manière à produire deux groupes de six diallèles (A et B) avec leurs réciproques. Les descendances étaient ensuite évaluées sur leur aptitude à la callogenèse. Des différences significatives (P < 0,01) étaient observées dans les deux diallèles pour les composantes croisement et aptitude générale à la combinaison (AGC). Les composantes aptitude spécifique à la combinaison (ASC) étaient significatives (P < 0,01), mais les croisement réciproque n'étaient pas significatifs (P < 0,05). Les valeurs d'héritabilité (h 2 ) pour la callogenèse par moyenne de parcelle étaient, respectivement, de 73 et de 61% pour les diallèles A et B. Comme ces coefficients sont élevés et vu l'absence d'effets significatifs pour les composantes croisement réciproque, les stratégies de sélection utilisant la sélection récur-rente devraient ouvrir la voie à la mise au point...

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Role of genetic background in somatic embryogenesis in Medicago

Cited by 152 publications

References 16 publications

Vicilin with carboxy‐terminal KDEL is retained in the endoplasmic reticulum and accumulates to high levels in the leaves of transgenic plants

Vicilin with carboxy‐terminal KDEL is retained in the endoplasmic reticulum and accumulates to high levels in the leaves of transgenic plants

Transient expression of a reporter gene changes significantly during somatic embryogenesis in alfalfa

Quantitative genetic analysis of in vitro callus proliferation in alfalfa

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