We have developed a viral RNA (vRNA) dot blot assay for identified a subset of candidate high-titer producer cell rapid identification of high-titer retrovirus vector production clones. In three independent screens the supernatant with by packaging cell clones. The procedure employs Trizol LS the highest biological titer was produced by one of the prereagent to purify vRNA from packaging cell supernatants, viously defined candidate high-titer producer clones. Our a sensitive dot blot assay, and PhosphorImager technology procedure greatly facilitates virus titration by: (1) rapidly elito quantify packaged viral genomes in 2 days. Experiments minating the vast majority of low-titer producer cell clones; performed on viral supernatants of known biological titer (2) accurately identifying the subset of candidate high-titer demonstrated that the vRNA dot blot assay was extremely producer clones for further biological titration and assesssensitive and that dot intensity correlated directly with viral ment of the proviral genomic structure; and (3) reducing titer. It is often necessary to analyze approximately 100 laborious tissue culture manipulations to a minimum. Furvirus producing cell clones, making this method useful as thermore, the reliance of this method on molecular deteca rapid screen to identify the highest virus producing tion makes it ideally suited for the isolation of high-titer clones. The vRNA dot blot assay consistently clones lacking a drug selection marker.

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