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After the first cycle of amplification, it is often possible to quantitate the cRNA yield at this stage. Routinely, 400 ng - 1 μg of cRNA can be obtained from 50 to 100 ng of starting material. However, it is difficult to use the cRNA yield as the only metric to determine the success of the assay. Therefore, it may be most informative to combine the results from the first-cycle IVT yield with the second-cycle IVT yield, as well as array-quality metrics and data on various positive controls, to obtain a comprehensive view of how the assay has proceeded.
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