A 153.2 kbp human bacterial artificial chromosome (BAC) CH523-111K12 (Chr9:27,561,112-27,714,301 : Hg19) was obtained containing the familial FTD/ALS human patient-derived C9orf72 gene. This genomic C9orf72 DNA fragment has 140.5 kbp of upstream sequences, the promoter region and exon 1 through part of exon 6 - including ~500 hexanucleotide repeat motifs (GGGGCC>[500]<) in the non-coding intron 1 and part of the 3' UTR of the short isoform V1. The BAC has 140.5 kbp of transcriptionally-upstream (telomeric) sequences reported to contain no other complete loci or confirmed genes. Southern blot analysis of hemizygous mice identified two transgene insertions - one with ~500 GGGGCC repeats and the other with ~300 GGGGCC repeats. The truncated human C9orf72 sequences in the transgene express three human C9ORF72 splice variant isoforms (V1, V2 and V3), of which transcripts V1 and V3 contain the hexanucleotide expansion. Human C9ORF72 mRNA transcripts were detected in all tissues examined. Human-specific probes targeting each variant mRNA detected all three transcripts at expression levels relative to their abundances in human frontal cortex. Moreover, the total level of the transgenic human transcripts was roughly comparable to that of the endogenous mouse C9ORF72 ortholog transcripts and to expression levels of human control and ALS/FTD cases. 3-5 copies of the transgene have integrated on chromosome 4 (23,004,513-23,004,540 [mouse mm10]) and resulted in a 27 bp deletion of genomic region in between the two identified breakpoints. No genes are annotated in this region. In addition, a total of 224 kb of E. coli sequence appears to be co-integrated after the transgene sequence. (J:228730)
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