The SAT line (Sanger AB Tübingen) is derived from two lines, AB and TU. A male doubled haploid AB (DHABC1b S#18732) and a female doubled haploid TU fish (DHTu2 S#18728) were generated by Amanda Rapp and Catherine Wilson in John Postlethwait’s lab. Both fish were sequenced using Illumina GA sequencing technology to ~40x coverage. Over 10 million SNPs, compared to Zv8, were mined from the sequences, and an Affymetrix custom SNP chip covering over 200,000 SNPs was designed (Item # 520747, Array Name ZFSNP200m520747F, Array Format: 49-7875). Each sequenced fish is homozygous, and was crossed to make genetically identical and heterozygous F1 hybrids: with one intact AB and TU chromosome from each parent. These F1s were crossed to make F2s, which should have one crossover per chromosome. This F2 cross was used to make the new SAT meiotic map of 140,306 SNPs across 430 F2s using MSTmap (M. D. Clark et al. unpublished). F2 adult pairs were shipped from the Sanger to ZIRC.
Due to the nature of this cross, we believe that the SAT line should be largely free of embryonic lethal mutations, because these should have been selected against by the doubled haploid process (any remaining lethal mutations might have been complemented by wild-type maternal mRNA or protein). Similarly trans-heterozygous lethal alleles should not exist in the F1s. For F2s and subsequent generations, the genome should be reconstructable by dense genotyping, e.g. using the 200k SNP array, calculating the crossovers and identifying the pieces of sequenced AB and TU genomes contained within a given fish. Thus, this strain is genetically defined, and the individual genomes are definable. Furthermore, it is largely free of lethal and deleterious alleles, and should possess hybrid vigor enabling use as a wild-type line and maintenance as a strain. We particularly recommend this line for use where the knowledge of exact genome sequence is important, e.g. morpholino and PCR primer design, and reverse genetics such as TILLING and zinc finger nuclease targeting.