Co-detection of chimeric bcr/abl (target) and β-actin (control) messenger RNA in individual CFU-GM colonies derived from CML patients using the polymerase chain reaction Academic Article Article uri icon

Overview

MeSH Major

  • Neoplasm Proteins
  • Neoplasms
  • Neovascularization, Pathologic
  • Proteome
  • Proteomics

abstract

  • In order to quantitate the magnitude of the normal and Philadelphia (Ph') chromosome-positive(+) progenitor cells for various research and clinical settings/studies, we have applied the highly sensitive polymerase chain reaction (PCR) for examining the cells contained in individual hematopoietic colonies for chimeric bcr/abl mRNA, a specific molecular marker for chronic myelogenous leukemia (CML). Thus, individual 14-day CFU-GM colonies, obtained by growth of bone marrow cells from CML patients were removed from methylcellulose cultures and total RNA from each colony was isolated. First-strand complementary DNAs (cDNA) corresponding to all mRNAs in the sample were obtained by using random hexamers in a reverse transcription (RT) reaction. cDNA then served as the substrate in the PCR. To ensure the integrity of the RNA extracted from each colony, beta-actin and bcr/abl cDNA sequences were amplified in the same reaction vessel. Using this method, we have examined the colonies grown from three CML patients and found that 5 out of 5, 9 out of 9 and 8 out of 9 colonies contained a bcr/abl transcript. This method is simple, highly sensitive and should facilitate studies comparing the expression of various oncogenes in normal and leukemic hematopoietic progenitor cells.

publication date

  • January 1991

Research

keywords

  • Academic Article

Identity

Digital Object Identifier (DOI)

  • 10.1016/0145-2126(91)90471-5

PubMed ID

  • 1921462

Additional Document Info

start page

  • 867

end page

  • 74

volume

  • 15

number

  • 9