Selecting somatic cell hybrids with hat media and nystatin methyl ester

Area of Science:

• Cell Biology
• Genetics
• Biochemistry

Background:

• Interspecific hybrid isolation requires selective methods to eliminate parental cells.
• Polyene antibiotics like nystatin methyl ester (NME) exhibit selective toxicity.
• Biochemically defective cell lines are crucial for genetic complementation studies.

Purpose of the Study:

• To evaluate nystatin methyl ester (NME) as a half-selection agent for isolating interspecific mouse-Syrian hamster hybrids.
• To demonstrate the efficiency of HAT-NME media in eliminating unfused parental cells.
• To confirm the expression of dominant polyene resistance in hybrid cells.

Main Methods:

• Utilized polyethylene glycol (PEG)-mediated fusion of biochemically defective mouse cells (A9 or B82) with genetically normal Syrian hamster cells (BHK/C13).
• Cultured fused cells in HAT media supplemented with NME (HAT-NME media).
• Monitored cell growth and isolated hybrid clones after 3 weeks.

Main Results:

• Unfused parental cells were selectively killed in HAT-NME media due to genetic defects (HGPRT- or TK-deficient) or NME sensitivity.
• Hybrid cells, exhibiting genetic complementation, proliferated successfully in HAT-NME media.
• Isolated hybrid clones demonstrated dominant expression of polyene resistance.

Conclusions:

• Nystatin methyl ester (NME) is an effective half-selection agent for isolating interspecific hybrids.
• The HAT-NME selection system efficiently eliminates unfused parental cells.
• This technique is valuable for generating hybrids between genetically defective and normal cells.