Vol 9 No1 - Journal of Cell and Molecular Biology - Haliç Üniversitesi

µl containing 50–100 ng DNA template in 10 mM
Tris–HCl (pH 8.0), 50 mM KCl, 1.5 mM MgCl2, 1
mM each of dNTPs, 1.0 U Taq DNA polymerase
(Fermentas, Germany) and 1.0 mM of each primer
previously described (Ye et al., 1996) The
conditions of PCR amplification were as follows:
denaturation step at 94˚C for 5 min followed by 35
cycles at 94˚C for 1 min, 65˚C for 45 s, 72˚C for 1
min, a final extension at 72˚C for 7 min and stop at
4˚C.
In order to complete genotype analyses, amplicons
MMP1 and MMP3 promotor polymorphisms in TMJ disorder 65
of MMP1 and MMP3 were digested by XmnI (New
England Biolabs) and Tth111I (New England
Biolabs) restriction enzymes, respectively, as
previously described (Planello et al., 2011). 1G
allele of MMP1 was digested to 89 and 29 bp
whereas 2G allele was not digested and remained in
its original amplicon length, 118 bp. 5A allele of
MMP3 yielded 97 and 33 bp products by digestion,
6A allele was not digested and maintained the
original length of 130 bp. All the visualization
processes were carried out by 12% polyacrylamide
gel stained with etidium bromide (Figure 1)
Figure 1. RFLP analyses of MMP1 and MMP3 genes, M: Molecular marker (Fermentas, Germany), 2G/2G:
Homozygous genotype of 2G allele for MMP1, 1G/2G: Heterozygous genotype for MMP1, 1G/1G:
Homozygous of 1G allele for MMP1, 5A/5A: Homozygous genotype of 5A allele for MMP3, 5A/6A:
Heterozygous genotype for MMP3, 6A/6A: Homozygous genotype of 6A allele for MMP3.
Statistical analysis
Statistical analysis for identifying the differences
between TMJ and control groups was performed
with Chi-square test by using SPSS 18.0. p0,05
was accepted as statistically significant whereas
values higher than 0,05 was not.
Results
For the MMP1 genotypes, 8 of the 35 patients and
11 of the 50 controls were 1G/1G, 14 patients and
28 were 1G/2G and 13 patients and 11 controls
were 2G/2G, respectively. Genotypes of MMP1
polymorphisms and p- values were summarized in
Table 1. When we compare two groups for MMP1
genotypes, we could not find any statistically
significant association, all of the p- values are
0,5%.
For the MMP3 genotypes, of the 50 controls, 8
were 5A/5A, 23 were 5A/6A and 19 were 6A/6A,
respectively. 6 of the 35 patients were 5A/5A, 14
were 5A/6A and 15 were 6A/6A. Genotypes of
MMP1 polymorphisms were summarized in Table
2. Like in MMP1 genotypes, we could not find any
statistically significant association in patient and
control group, the p- values are 0,5%.