Cloning of the 5′ mRNA for the 230-kD bullous pemphigoid antigen by rapid amplification of cDNA ends

George Elgart, John R. Stanley

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

The 230-kD bullous pemphigoid antigen (BPAG1), defined by autoantibodies in patient sera, is a hemidesmosomal plaque protein in the same gene family as the intracellular proteins desmoplakin I/II and plectin. We had previously isolated, from a λgt11 library, overlapping cDNA clones with 6921 by of mRNA sequence for BPAG1. The coding sequence encoded by these clones included the 3′ stop codon but not the 5′ coding and non-coding region of the mRNA. To obtain these sequences we used the polymerase chain reaction (PCR) method called rapid amplification of cDNA ends (RACE). The PCR products were cloned into plasmids and sequenced. With five PCR primers we were able to obtain overlapping clones containing the 5′ region of the mRNA. An upstream stop codon in frame with the rest of the coding sequence demonstrates that the full 5′ coding sequence is obtained. Four different PCR products from two separate reactions had the same 5′ end, suggesting that this 5′ end is near, or at, the transcription start site. No alternatively spliced clones were found and no transmembrane site was predicted, confirming that BPAG1 is an intracellular hemidesmosomal plaque protein.

Original languageEnglish
Pages (from-to)244-246
Number of pages3
JournalJournal of Investigative Dermatology
Volume101
Issue number2
StatePublished - Aug 1 1993
Externally publishedYes

Fingerprint

Bullous Pemphigoid
Cloning
Polymerase chain reaction
Amplification
Organism Cloning
Complementary DNA
Clone Cells
Desmoplakins
Antigens
Polymerase Chain Reaction
Messenger RNA
Terminator Codon
Reaction products
Plectin
Proteins
Transcription Initiation Site
Gene Library
Autoantibodies
Plasmids
Genes

Keywords

  • adhesion
  • desmoplakin
  • hemidesmosome

ASJC Scopus subject areas

  • Dermatology

Cite this

Cloning of the 5′ mRNA for the 230-kD bullous pemphigoid antigen by rapid amplification of cDNA ends. / Elgart, George; Stanley, John R.

In: Journal of Investigative Dermatology, Vol. 101, No. 2, 01.08.1993, p. 244-246.

Research output: Contribution to journalArticle

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