qRTPCR was used to screen the line for Pseudoloma neurophilia. Each new generation is generated from the offspring of at least 25 small group crossings, each of which contains three males and three females. To produce the following generation, an equal number of donors are drawn from each cross. Embryos are examined for morphological defects on a daily basis until 5 days after fertilisation.

ENU Mutagenesis in Zebrafish –

ENU (N-ethyl-N-nitrosourea) mutagenesis is a well-known and well-proven technique for introducing random point mutations into the genome. Because there are presently no targeted knockout techniques for zebrafish, random mutagenesis is the favoured strategy in both forward and reverse genetic approaches. To generate high-density mutagenized zebrafish, six successive ENU treatments are given to adult male zebrafish at weekly intervals by immersing them in ENU solution. typically achieve an average germ line mutation burden of one mutation per 1.0 x 10(5)-1.5 x 10(5) basepairs using this method.