High efficiency DNA-mediated transformation of primate cells

Area of Science:

• Molecular Biology
• Cell Biology
• Genetics

Background:

• Mammalian cell lines are crucial for studying gene function and disease.
• Efficient gene transfer methods are essential for genetic manipulation in mammalian cells.
• Dominant selectable markers facilitate the identification of successfully transfected cells.

Purpose of the Study:

• To assess the stable transformation efficiency of various mammalian cell lines using dominant selectable markers.
• To compare the transformation frequencies across different species and cell types, including primate lines.
• To identify cell lines with high transfection efficiency for potential use in genetic research.

Main Methods:

• Recombinant vectors encoding Escherichia coli xanthine-guanine phosphoribosyltransferase (HPRT) or Tn5 aminoglycoside phosphotransferase (APH) were constructed.
• These vectors were introduced into several mammalian cell lines, including human HeLa, SV40-transformed xeroderma pigmentosum, and African green monkey kidney CV-1 cells.
• Stable transformation frequencies were determined by selecting for cells expressing the selectable markers.

Main Results:

• Human HeLa and SV40-transformed xeroderma pigmentosum cells achieved stable transformation frequencies of at least 10(-3) (0.1%).
• The CV-1 African green monkey kidney cell line demonstrated a significantly higher stable transformation frequency, reaching 6 X 10(-2) (6%).
• These results indicate variability in transfection efficiency among different mammalian cell types.

Conclusions:

• Mammalian cell lines can be stably transformed using dominant selectable markers like HPRT and APH.
• The CV-1 cell line exhibits exceptional efficiency for stable transformation, making it a valuable model for genetic studies.
• The findings highlight the importance of cell line selection for optimizing gene transfer experiments in mammalian systems.