Supplementary Table 2.
The primer sets for genes related to ossification used in RT-PCR
| Genes | Primer sets |
|---|---|
| DMP1 | 5’AGTGCCCAAGATACCACCAG3’ (sense) |
| 5’CATTCCCTCATCGTCCAACT3’ (antisense) | |
| RUNX2 | 5’GAATGCTTCATTCGCCTCACA3’ (sense) |
| 5’TGACCTGCGGAGATTAACCAT3’ (antisense) | |
| OCN | 5’AAGAGACCAAGGCGCTACCT3’ (sense) |
| 5’GCCGATAGGCCTCCTTGAAAG3’ (antisense) | |
| BMP2 | 5’CCCCTACATGCTAGACCTGTATCG3’ (sense) |
| 5’CACTCGTTTCTGGTAGTTCTTCCA’ (antisense) | |
| BMP6 | 5’CGGGTCTCCAGTGCTTCAGATT3’ (sense) |
| 5’GCATCCACAAGCTCTTACAACC3’ (antisense) | |
| GAPDH | 5’TGAACGGGAAGCTCACTGG3’ (sense) |
| 5’TCCACCACCCTGTTGCTGTA3’ (antisense) |
In the discovery cohort, we found an association between RUNX2, BMP2, and BMP6 and AS. We explored whether the DMP1 gene promoted the expression of RUNX2, BMP2, and BMP6 genes in transfected U2OS cells to examine the downstream pathways of DMP1 and to clarify the ectopic ossification mechanisms of AS. We detected the mRNA expression of OCN in transfected U2OS cells because the OCN protein is a marker of the bone formation process. Many studies use OCN as a preliminary biomarker of the efficacy of a given drug on bone formation. RT–PCR: Reverse transcription–polymerase chain reaction; DMP1: Dentin matrix protein 1; RUNX2: Runtrelated transcription factor 2; OCN: Osteocalcin; GAPDH: Glyceraldehyde–3–phosphate dehydrogenase; mRNA: MessengerRNA.