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At least 631 records · Page 35Linked to original sources
PubMed1985

Complementary DNA cloning and expression of the papaya ringspot potyvirus sequences encoding capsid protein and a nuclear inclusion-like protein in Escherichia coli.

Three cDNA clones that express viral gene products in Escherichia coli JM83 were derived from a watermelon mosaic virus-1 strain of papaya ringspot virus (PRSV-W). DNAs complementary to portions of the viral RNA were inserted into the pUC8 and pUC9 plasmids, and the expressed polypeptides were fusion products with the amino terminus of beta-galactosidase. Clones W1-77 and W2-1 expressed fusion products with apparent molecular weights of 40,000 (40K) and 14K, respectively, which were serologically related to PRSV capsid protein. A 52K product serologically related to a 54K nuclear inclusion protein of tobacco etch virus was produced by clone W1-18. The sequences encoding the capsid and 57K nuclear inclusion-like proteins of PRSV were physically mapped to adjacent positions through Southern blot analyses of clones W1-77 and W1-18.

Capsid↗
PubMed1988

Isolation of complementary DNA for bullous pemphigoid antigen by use of patients' autoantibodies.

Autoantibodies from bullous pemphigoid (BP) patients define a 230-kD protein found in the basement membrane of stratified squamous epithelia. The purpose of this study was to isolate and characterize a cDNA clone with coding sequences for BP antigen. Poly(A+) RNA derived from total RNA of cultured keratinocytes was used, with oligo-dT priming, to construct a cDNA library in the lambda gt11 expression vector, which was screened by the immunoperoxidase method with one BP serum. One darkly stained clone, called here the BP clone, was further characterized. 9 of 9 BP sera, but none of 6 normal and 11 pemphigus sera, bound the plaques of this BP clone. Furthermore, BP IgG affinity purified on plaques of this clone, but not unrelated clones, bound the epidermal basement membrane by immunofluorescence and immunoprecipitated the 230-kD BP antigen from extracts of cultured keratinocytes. Eco RI digestion of the BP clone's cDNA insert demonstrated a 680- and 1,500-bp fragment. Northern blots of total keratinocyte RNA showed that complementary riboprobes transcribed from both fragments hybridized to a 9-kb RNA. Dideoxy DNA sequencing from the 5' end of the BP cDNA demonstrated a 1,992-bp open reading frame, encoding a peptide of 76 kD. This BP cDNA clone will be valuable for understanding the protein structure, expression, and gene organization of BP antigen.

Amino Acid Sequence↗
PubMedDate not supplied

[Synthesis and cloning of DNA, complementary to rabbit globin pre-mRNA].

cDNA synthesized on rabbit bone marrow erythroid cells pre-mRNA was cloned in bacterial plasmids. Cold phenol extracted pre-mRNA was a several times more effective template in the reaction of reverse transcription without oligo(dT) 10-primer than hot phenol extracted pre-mRNA. There was no yield increase of DNA-product on hot phenol extracted pre-mRNA in the reaction of reverse transcription with the oligo(dT)10-primer addition. The "hot phenol" poly (A)+-pre-mRNA was used to obtain the representative, full-sized cDNA. The double-stranded form of this cDNA, obtained with the help of DNA-polymerase I, was inserted into the PstI-site of pBR322 plasmid. About 25% E. coli JC5183 clones, transformed with this hybrid plasmid, were found to contain the globin sequences.

Animals↗