Purification of the mRNA for chicken very low density lipoproteirtIIand molecular cloning of its full-length double-stranded cDNA | Zendy

Bé Wieringa | Zendy; Willem G. Roskam | Zendy; Annika C. Arnberg | Zendy; Janneke van der Zwaag-Gerritsen | Zendy; Geert AB | Zendy; Max Gruber | Zendy

Open Access

Purification of the mRNA for chicken very low density lipoproteirt_IIand molecular cloning of its full-length double-stranded cDNA

Author(s) -

Bé Wieringa,

Willem G. Roskam,

Annika C. Arnberg,

Janneke van der Zwaag-Gerritsen,

Geert AB,

Max Gruber

Publication year - 1979

Publication title -

nucleic acids research

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 9.008

H-Index - 537

eISSN - 1362-4954

pISSN - 0305-1048

DOI - 10.1093/nar/7.8.2147

Subject(s) - biology , complementary dna , microbiology and biotechnology , recombinant dna , reverse transcriptase , molecular cloning , plasmid , restriction enzyme , messenger rna , gel electrophoresis , nucleic acid thermodynamics , restriction map , dna , oligonucleotide , rna , gene , biochemistry

The mRNA coding for the small apo-Very Low Density Lipoprotein (apo-VLDLII) from chicken serum was highly enriched by oligo(dT) chromatography and preparative gel electrophoresis of estrogenised liver RNA. Double-stranded cDNA was synthesised by the subsequent actions of reverse transcriptase and DNA polymerase, and used for a preliminary characterisation of the structural gene. Molecular cloning of dC-tailed ds-cDNA into the Pst I site of plasmid pBR 322 yielded several recombinant clones. Five chimeric DNAs were selected and characterised by restriction enzyme mapping and electron microscopy of R-loops. At least two of them (pVLDLII 3.33 and pVLDLII 4.82) contain an almost full-length ds-transcript of VLDLII mRNA in which no more than 10-20 bases at the 5'- end are missing.

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