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. 2015 Jun 4;10(6):e0128275. doi: 10.1371/journal.pone.0128275

Fig 1. OC-STAMP knockout allele, radiographs.

Fig 1

A. Gene targeting vector is a “knockout first” homologous recombination gene trap with a selection cassette and recombination sites for Cre (loxP) and flippase (Frt), as indicated. This allele, which results in a complete loss-of-function, is designated Ocstamp tm1a(KOMP)Wtsi. Exons in the upper (gene) and lower (targeting construct) diagrams are numbered 1, 2, and 3. The selection cassette in intron 1 eliminates production of any functional OC-STAMP. The 5’ and 3’ UTR’s are shown in boxes, and the 3’ alternative end is shown in light blue. Asterisks indicate locations of genotyping primers. From left to right they are: OC-F1 (red) (5’-TTGCCTGTAAATGATGGAGTGGGC-3’); En2R1 (blue) (5’-TGGTGTGGGAAAGGGTTCGAAGTT-3’); OCR1E (purple) (5’-TGGCGCAGCTGGTAAGTGGTATTA-3’). These give PCR products of 1044 bp from WT (OCF1 and OCR1E) and 297 bp from the knockout allele (OCF1 and En2R1). To confirm correct 3’ end insertion, the right-hand pair of primers are: LoxPF (green) (5’-GAGATGGCGCAACGCAATTAAT-3’) and SR1 (brown) (5’-CTGTGACTAAGTAACCATCAAAGCGG-3’), which give a 681 bp product only in the targeted allele (not shown). B. PCR of genomic DNA from the mice yielded the expected sizes in homozygous wild type (+/+; upper arrowhead indicates 1044 bp), heterozygotes (+/-), and homozygous knockout (-/-; lower arrowhead indicates 297 bp). C. X-rays of 2-week-old wild type (left) and OCSt-KO mice showed no differences in either whole-body views (upper) or rear quadrant views (lower panels). All skeletal elements appear normally formed and without accumulation of trabecular or cortical bone.