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Figure 7. Reverse Screen Strategy ENU-treated sperm (G0) was used to fertilize wild-type eggs (in vitro fertilization), and the resulting F1 families raised to adulthood. F1 males were killed and their testes dissociated, with a portion used to generate F2 tadpoles and the remainder frozen in several aliquots per individual (F1 library). F1 females were used in the gynogenetic forward screens (see Figure 1). F2 genomic DNA was isolated from the tadpoles for reverse genetic (TILLING) screens. Known genomic sequences were used to design nested PCR primers, and then individual F2 tadpole amplicons were sequenced to detect induced mutations. Mutations are then recovered from frozen testes by in vitro fertilization for subsequent phenotypic analysis. doi:10.1371/journal.pgen.0020091.g007 Image published in: Goda T et al. (2006) Copyright: © 2006 Goda et al. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution license Larger Image Printer Friendly View

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