In vivo expression of a full-length cDNA copy of potato leafroll virus (PLRV) in protoplasts and transgenic plants

1997 ◽  
Vol 253 (5) ◽  
pp. 609-614 ◽  
Author(s):  
D. Prüfer ◽  
J. Schmitz ◽  
E. Tacke ◽  
B. Kull ◽  
W. Rohde
Keyword(s):  
Transgenic Plants ◽  
Potato Leafroll Virus ◽  
Full Length ◽  
Full Length Cdna ◽  
Cdna Copy ◽  
In Vivo Expression ◽  
Leafroll Virus

Establishment of Full-length cDNA Clones and an Efficient Oral Infection Model for Feline Coronavirus in Cats

2021 ◽  
Author(s):  
Gang Wang ◽  
Guangli Hu ◽  
Rui Liang ◽  
Jiale Shi ◽  
Xiuxiu Qiu ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Full Length ◽  
Infection Model ◽  
Type I ◽  
Feline Infectious Peritonitis ◽  
Spike Gene ◽  
Full Length Cdna ◽  
Adult Cats

Feline infectious peritonitis virus (FIPV) is the etiologic agent of feline infectious peritonitis (FIP) and causes fatal disease in cats of almost all ages. Currently, there are no clinically approved drugs or effective vaccines for FIP. Furthermore, the pathogenesis of FIP is still not fully understood. There is an urgent need for an effective infection model of feline infectious peritonitis induced by FIPV. Here, we constructed a field type I FIPV full-length cDNA clone, pBAC-QS, corresponding to the isolated FIPV QS. By replacing the FIPV QS spike gene with the commercially available type II FIPV 79-1146 (79-1146_CA) spike gene, we established and rescued a recombinant virus, designated rQS-79. Moreover, we constructed 79-1146_CA infectious full-length cDNA pBAC-79-1146_CA, corresponding to recombinant FCoV 79-1146_CA (r79-1146_CA). In animal experiments with one- to two-year-old adult cats orally infected with the recombinant virus, rQS-79 induced typical FIP signs and 100% mortality. In contrast to cats infected with rQS-79, cats infected with 79-1146_CA did not show obvious signs. Furthermore, by rechallenging rQS-79 in surviving cats previously infected with 79-1146_CA, we found that there was no protection against rQS-79 with different titers of neutralizing antibodies. However, high titers of neutralizing antibodies may help prolong the cat survival time. Overall, we report the first reverse genetics of virulent recombinant FCoV (causing 100% mortality in adult cats) and attenuated FCoV (causing no mortality in adult cats), which will be powerful tools to study the pathogenesis, antiviral drugs and vaccines for FCoV. Importance Tissue- or cell culture-adapted feline infectious peritonitis virus (FIPV) usually loses pathogenicity. To develop a highly virulent FIPV, we constructed a field isolate type I FIPV full-length clone with the spike gene replaced by the 79-1146 spike gene, corresponding to a virus named rQS-79, which induces high mortality in adult cats. rQS-79 represents the first described reverse genetics system for highly pathogenic FCoV. By further constructing the cell culture-adapted FCoV 79-1146_CA, we obtained infectious clones of virulent and attenuated FCoV. By in vitro and in vivo experiments, we established a model that can serve to study the pathogenic mechanisms of FIPV. Importantly, the wild-type FIPV replicase skeleton of serotype I will greatly facilitate the screening of antiviral drugs, both in vivo and in vitro.

In Vivo Expression of Full-Length Human Dystrophin from Adenoviral Vectors Deleted of All Viral Genes

1996 ◽  
Vol 7 (15) ◽  
pp. 1907-1914 ◽  
Author(s):  
Sarah Ehlen Haecker ◽  
Hansell H. Stedman ◽  
Rita J. Balice-Gordon ◽  
Daniel B. J. Smith ◽  
James P. Greelish ◽  
...  
Keyword(s):  
Adenoviral Vectors ◽  
Full Length ◽  
Viral Genes ◽  
In Vivo Expression ◽  
Human Dystrophin

scholarly journals Isolation and functional analysis of a cDNA for human Jagged2, a gene encoding a ligand for the Notch1 receptor.

1997 ◽  
Vol 17 (10) ◽  
pp. 6057-6067 ◽  
Author(s):  
B Luo ◽  
J C Aster ◽  
R P Hasserjian ◽  
F Kuo ◽  
J Sklar
Keyword(s):  
Feedback Control ◽  
Expressed Sequence Tag ◽  
Myogenic Differentiation ◽  
Notch Receptor ◽  
Full Length ◽  
C2c12 Cells ◽  
Cdna Clones ◽  
Gene Encoding ◽  
Full Length Cdna

Signaling through Notch receptors has been implicated in the control of cellular differentiation in animals ranging from nematodes to humans. Starting from a human expressed sequence tag-containing sequence resembling that of Serrate, the gene for a ligand of Drosophila melanogaster Notch, we assembled a full-length cDNA, now called human Jagged2, from overlapping cDNA clones. The full-length cDNA encodes a polypeptide having extensive sequence homology to Serrate (40.6% identity and 58.7% similarity) and even greater homology to several putative mammalian Notch ligands that have subsequently been described. When in situ hybridization was performed, expression of the murine Jagged2 homolog was found to be highest in fetal thymus, epidermis, foregut, dorsal root ganglia, and inner ear. In Northern blot analysis of RNA from tissues of 2-week-old mice, the 5.0-kb Jagged2 transcript was most abundant in heart, lung, thymus, skeletal muscle, brain, and testis. Immunohistochemistry revealed coexpression of Jagged2 and Notch1 within thymus and other fetal murine tissues, consistent with interaction of the two proteins in vivo. Coculture of fibroblasts expressing human Jagged2 with murine C2C12 myoblasts inhibited myogenic differentiation, accompanied by increased Notch1 and the appearance of a novel 115-kDa Notch1 fragment. Exposure of C2C12 cells to Jagged2 led to increased amounts of Notch mRNA as well as mRNAs for a second Notch receptor, Notch3, and a second Notch ligand, Jagged1. Constitutively active forms of Notchl in C2C12 cells also induced increased levels of the same set of mRNAs, suggesting positive feedback control of these genes initiated by binding of Jagged2 to Notch1. This feedback control may function in vivo to coordinate differentiation across certain groups of progenitor cells adopting identical cell fates.

scholarly journals Evidence for RNA-mediated defence effects on the accumulation of Potato leafroll virus

2001 ◽  
Vol 82 (12) ◽  
pp. 3099-3106 ◽  
Author(s):  
Hugh Barker ◽  
Kara D. McGeachy ◽  
Eugene V. Ryabov ◽  
Uli Commandeur ◽  
Mike A. Mayo ◽  
...  
Keyword(s):  
Coat Protein ◽  
Transgenic Plants ◽  
Vascular Tissue ◽  
Fluorescent Protein ◽  
Vascular System ◽  
Potato Leafroll Virus ◽  
Transcriptional Gene Silencing ◽  
Mesophyll Cells ◽  
Tobacco Plants ◽  
Leafroll Virus

In plants infected with Potato leafroll virus (PLRV), or other luteoviruses, infection is very largely confined to cells in the vascular system. Even in tobacco plants transformed with PLRV full-length cDNA, in which all mesophyll cells should synthesize infectious PLRV RNA transcripts, only a minority of the mesophyll cells accumulate detectable amounts of virus. We have explored this phenomenon further by transforming a better PLRV host, Nicotiana benthamiana, with the same transgene, by superinfecting transformed plants with Potato virus Y and by producing tobacco plants in which cells contained both PLRV cDNA and DNA encoding the P1/HC-Pro genes of the potyvirus Tobacco etch virus. A greater proportion of cells in superinfected plants or in doubly transgenic plants accumulated PLRV than did in singly transgenic tobacco plants. However, most cells in these plants did not accumulate virus. To investigate restriction of the multiplication of viruses containing PLRV sequences, transgenic plants were infected with a chimeric virus that consisted of Tobacco mosaic virus (TMV) containing genes for either the coat protein (CP) of PLRV or jellyfish green fluorescent protein (GFP) in place of the TMV coat protein. The virus that encoded PLRV CP spread more slowly and accumulated less extensively than did the virus that expressed GFP. The results support the suggestion that an RNA-mediated form of resistance that resembles post-transcriptional gene silencing operates in non-vascular cells and may be part of the mechanism that restricts PLRV to vascular tissue in conventionally infected plants.

A fully recombinant ELISA using in vivo biotinylated antibody fragments for the detection of potato leafroll virus

2009 ◽  
Vol 159 (2) ◽  
pp. 200-205 ◽  
Author(s):  
Ahmad Al-Mrabeh ◽  
Angelika Ziegler ◽  
Graham Cowan ◽  
Lesley Torrance
Keyword(s):  
Potato Leafroll Virus ◽  
Antibody Fragments ◽  
Leafroll Virus

scholarly journals In vivo evidence for posttranslational translocation and signal cleavage of the killer preprotoxin of Saccharomyces cerevisiae.

1986 ◽  
Vol 6 (12) ◽  
pp. 4274-4280 ◽  
Author(s):  
S J Lolle ◽  
H Bussey
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasmid Vector ◽  
Full Length ◽  
Protein Species ◽  
Coding Region ◽  
Double Stranded Rna ◽  
Full Length Cdna ◽  
Electrophoretic Mobilities ◽  
K1 Toxin

A full-length cDNA of the M1 double-stranded RNA killer preprotoxin coding region successfully directed the synthesis of secreted K1 toxin when expressed in Saccharomyces cerevisiae from a plasmid vector. Three protein species immunoreactive with antitoxin antiserum were detected intracellularly in transformants harboring this killer cDNA plasmid. These toxin precursor species were characterized by using secretory-defective hosts, by comparative electrophoretic mobilities, and by tunicamycin susceptibility. Such studies indicate that these three protein species represent intermediates generated by signal cleavage of the preprotoxin and its subsequent glycosylation and provide evidence that these events occur posttranslationally.

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