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phosphoribosyl transferase

Bromodeoxyuridine

Thymidine kinase

possible to select against the growth of these enzvme-deficient plasmacytoma lines by forcing them to use the salvage pathway for DNA synthesis. This is achieved by using the folic acid analog, aminopterin, which blocks the normal pathway for purine and pyrimidine synthesis, as shown in Figure 2. The plasmacytoma is then forced to use the salvage pathway but because the mutants lack one of the necessary enzymes they do not survive in the HAT medium.

The genetic defect in such mutant plasmacytoma lines that results in the absence of a functional salvage pathway is of little importance during growth of these cells in normal medium because these enzymes are involved only in salvage pathways synthesis, and under normal conditions this alternative route of nucleotide synthesis is unnecessary. Spleen cells, on the other hand, can synthesize a complete set of salvage pathway enzymes and, when fused to the HGPRT or TK plasmacytoma cells during hybrid formation, enable the hybrid cells to produce sufficient HGPRT or TK for the fused cells to survive in the HT medium which contains hypoxanthine and thymidine. In this way, nonfused plasmacytoma cells are killed, while the hybrids resulting from fusions with normal cells continue to grow. In contrast, normal unfused B cells have only a limited lifespan in vitro because they are not transformed, do not proliferate and eventually die.

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