Transfer and expression of the bacterial NPT-II gene In chick embryos using a Schmidt-Ruppin retrovirus vector

Mature pollen of lodgepole pine (Pinusconcorta Dougl.), yellow cypress (Chamaecyparisnootkatensis (D. Don) Spach), western hemlock (Tsugaheterophylla (Raf.) Sarg.), jack pine (Pinusbanksiana Lamb.), and black spruce (Piceamariana (Mill.) B.S.P.) was bombarded with gold particles coated with four different plasmid constructions, pRT99GUS, pBM113Kp, pAct1-D, and pGA984, using the biolistic PDS-1000/He device. A protocol was devised for efficient gene transfer and gene expression assay in pollen. False positive results for expression of the β-glucuronidase (GUS) gene assayed with the substrate X-glucuronide were observed with pollen of yellow cypress, western hemlock, and lodgepole pine. The highest levels of transient GUS gene expression were obtained with plasmid pBM113Kp, which carried the GUS gene under the control of the wheat abscisic acid inducible early methionine promoter. The plasmids pRT99GUS (35S promoter) and pAct1-D (rice actin promoter) yielded similar intermediate levels of transient GUS gene expression. The pollen-specific promoter of the α-tubulin gene from Arabidopsisthaliana (pGA984) yielded the lowest levels of gene expression in pollen. Of the four species, yellow cypress showed the lowest levels of transient GUS gene expression and black spruce yielded the highest levels. The neomycin phosphotransferase II (NPT II) gene was also tested as a reporter gene for pollen transformation and was easily assayed via ELISA. The fusion gene between NPT II and GUS genes was detected at a lower level than the nonfused NPT II gene when under the control of the same 35S promoter. The method devised here could be used for the study of tissue-specific gene expression in conifer pollen.

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Transfer of Lycopersicon chilense Chitinase Gene to Cultivated Tomato (L. esculentum)

HortScience ◽

10.21273/hortsci.30.4.878g ◽

1995 ◽

Vol 30 (4) ◽

pp. 878G-878

Author(s):

Zahra Agharbaoui ◽

Long Xi Yu ◽

Viano Poysa ◽

Zohreh Tabaeizadeh

Keyword(s):

Chitinase Gene ◽

Indirect Organogenesis ◽

Tomato Cultivar ◽

Wild Tomato Species ◽

Lycopersicon Chilense ◽

Tomato Species ◽

Npt Ii ◽

Selection Medium ◽

Polymerase Chain ◽

Npt Ii Gene

We have isolated a drought-induced chitinase gene from L. chilense, a wild tomato species. Owing to our interests in genetic improvement of cultivated tomato, we have transferred the L. chilense chitinase gene to this species. The transformation plasmid constructed contained the coding sequence of L. chilense chitinase gene linked to CaMVS35 promoter as well as the NPT II gene linked to nopaline synthetase promoter. The leaf disk transformation regeneration technique was applied to one commercial tomato cultivar and four inbred lines. Shoots were produced on the selection medium through direct or indirect organogenesis. Plantlets that have been rooted on kanamycin-containing medium were transferred to soil where they grew to maturity and produced flowers and fruit. The transgenic nature of some of the analyzed plants was confirmed by polymerase chain reaction. Research is continuing to evaluate transgenic plants with regard to their level of tolerance to phytogenic fungi.

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Identification of plant promoters in situ by T-DNA-mediated transcriptional fusions to the npt -II gene

The EMBO Journal ◽

10.1002/j.1460-2075.1986.tb04423.x ◽

1986 ◽

Vol 5 (8) ◽

pp. 1755-1760 ◽

Cited By ~ 62

Author(s):

T.H. Teeri ◽

L. Herrera-Estrella ◽

A. Depicker ◽

M. Van Montagu ◽

E.T. Palva

Keyword(s):

Npt Ii ◽

Plant Promoters ◽

Npt Ii Gene ◽

Transcriptional Fusions

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Agrobacterium-Mediated Transformation of Three Freshwater Microalgal Strains

Polish Journal of Microbiology ◽

10.33073/pjm-2014-052 ◽

2014 ◽

Vol 63 (4) ◽

pp. 387-392 ◽

Cited By ~ 6

Author(s):

MARY SANITHA ◽

SUDHAKAR RADHA ◽

ANWAR ALIYA FATIMA ◽

SELVARAJU GAYATHRI DEVI ◽

MOHANDASS RAMYA

Keyword(s):

Morphological Characteristics ◽

Pcr Amplification ◽

Freshwater Environment ◽

Agrobacterium Mediated Transformation ◽

Chlorella Sp ◽

Gus Histochemical Assay ◽

Npt Ii ◽

Scenedesmus Bajacalifornicus ◽

Npt Ii Gene ◽

Inexpensive Procedure

Microalgal transformation has gained interest in recent years. Agrobacterium-mediated transformation remains as the most efficient method for the development of transgenic plants and microalgae due to its wide host range, inexpensive procedure and transfer of large segments of DNA. In the present study, three different microalgal species were isolated from freshwater environment and identified based on the morphological characteristics and ITS-2 region amplification. Agrobacterium-mediated transformation was successful for the isolates Chlorella sp., Ankistrodesmus sp and Scenedesmus bajacalifornicus. Gene integration and expression was confirmed by PCR amplification of hptII and GUS histochemical assay. A. tumifaciens contamination was checked by amplification of npt II gene (kanamycin resistant) which lies outside the T-border. Based on GUS assay, transformation efficiencies were found to be 12.25% for Chlorella sp. 2.96% for Scenedesmus bajacalifornicus and 3.5% for Ankistrodesmus sp.

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Dexamethasone Effect on Embryonic Chick Respiratory Epithelium

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100053905 ◽

1982 ◽

Vol 40 ◽

pp. 248-249

Author(s):

M.R. Richter ◽

R.V. Blystone

Keyword(s):

Lung Development ◽

Critical Role ◽

Respiratory Epithelium ◽

Chick Embryos ◽

Embryonic Chick ◽

White Leghorn ◽

Lung Maturation ◽

Synthetic Analogs ◽

Lung Physiology ◽

Avian Lung

Dexamethasone and other synthetic analogs of corticosteroids have been employed clinically as enhancers of lung development. The mechanism(s) by which this steroid induction of later lung maturation operates is not clear. This study reports the effect on lung epithelia of dexamethasone administered at different intervals during development. White Leghorn chick embryos were used so as to remove possible maternal and placental influences on the exogenously applied steroid. Avian lung architecture does vary from mammals; however, respiratory surfactant produced by the lung epithelia serves an equally critical role in avian lung physiology.

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Development of the Foramen Secundum in the Chick as Observed by Scanning Electron Microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100091044 ◽

1976 ◽

Vol 34 ◽

pp. 212-213

Author(s):

M.J.C. Hendrix ◽

D.E. Morse

Keyword(s):

Electron Microscopy ◽

Scanning Electron Microscopy ◽

Procaine Hydrochloride ◽

Cardiac Anomaly ◽

Chick Embryos ◽

Congenital Cardiac Anomaly ◽

Septal Defects ◽

Critical Point Drying ◽

Scanning Electron

Atrial septal defects are considered the most common congenital cardiac anomaly occurring in humans. In studying the normal sequential development of the atrial septum, chick embryos of the White Leghorn strain were prepared for scanning electron microscopy and the results were then extrapolated to the human heart. One-hundred-eighty chick embryos from 2 to 21 days of age were removed from their shells and immersed in cold cacodylate-buffered aldehyde fixative . Twenty-four embryos through the first week post-hatching were perfused in vivo using cold cacodylate-buffered aldehyde fixative with procaine hydrochloride. The hearts were immediately dissected free and remained in the fixative a minimum of 2 hours. In most cases, the lateral atrial walls were removed during this period. The tissues were then dehydrated using a series of ascending grades of ethanol; final dehydration of the tissues was achieved via the critical point drying method followed by sputter-coating with goldpalladium.

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Immunocytochemical localization of desmin and vimentin in chick embryonic myocardial cells

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100159783 ◽

1990 ◽

Vol 48 (3) ◽

pp. 448-449

Author(s):

Yukiko Sugi

Keyword(s):

Sodium Methoxide ◽

Intermediate Filament Protein ◽

Chick Embryos ◽

Immunocytochemical Localization ◽

Myocardial Cells ◽

Immunofluorescence Study ◽

Immunofluorescent Localization ◽

Rabbit Igg ◽

Semithin Sections ◽

Filament Protein

In cultured skeletal muscle cells of chick, one intermediate filament protein, vimentin, is primarily formed and then synthesis of desmin follows. Coexistence of vimentin and desmin has been immunocytochemically confirmed in chick embryonic skeletal musclecells. Immunofluorescent localization of vimentin and desmin has been described in developing myocardial cells of hamster. However, initial localization of desmin and vimentin in early embryonic heart has not been reported in detail. By quick-freeze deep-etch method a loose network of intermediate filaments was revealed to exist surrounding myofibrils. In this report, immunocytochemical localization of desmin and vimentin is visualized in early stages of chick embryonic my ocardium.Chick embryos, Hamburger-Hamilton (H-H) stage 8 to hatch, and 1 day old postnatal chicks were used in this study. For immunofluorescence study, each embryo was fixed with 4% paraformaldehyde and embedded in Epon 812. De-epoxinized with sodium methoxide, semithin sections were stained with primary antibodies (rabbit anti-desmin antibody and anti-vimentin antibody)and secondary antibody (RITC conjugated goat-anti rabbit IgG).

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