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Lack of Rpc9 is responsible for the phenotype of mutant line 116 embryos. (A) Schematic of positional cloning. a-g indicate the locations of polymorphic primers distributed between 29,708,538 to 30,321,343bp of chromosome 21 and the ratios of recombination out of total 2208 embryos in each position. Genes scattered in this region are marked and rpc9 (in red) is adjacent to the polymorphic marker d. Note that the chromosomal locations and sequences of these primers are listed in Table S3. (B) Gene sequencing identified a transversion of T>A in the second exon (exon 2) of rpc9, which formed a pre-mature stop code in the coding region of rpc9 in mutant line 116 embryos. (C) Western blot showed that the protein level of Rpc9 was greatly decreased in mutants at 4dpf. (D) Injection of the rpc9 overexpression plasmid driven by hsp70 promoter (hsp70-rpc9-egfp) exerted a full rescue effect of rag1. Red arrowheads in D indicate the thymus.
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