Probes were purchased from Invitrogen Corp (Invitrogen, California, USA). The thermal cycling conditions were: 5 minutes at 95°C, followed by 60 cycles of 30 seconds at 95°C and 1 minute at 60°C. Statistic analysis The Levene’s test was performed to determine the homogeneity of variance for all the data, and then the paired Chi-Square test or 2-related samples Wilcoxon nonparametric test was performed to #selleck screening library randurls[1|1|,|CHEM1|]# compare the positive rates and the levels of DHX32 gene expression between tumor tissue and its adjacent normal tissue
in each patient, respectively. The Mann-Whitney U test was used for comparing the gene expression of DHX32 between the different groups according to various clinical and pathological variables. All of the statistical Milciclib mouse analyses were performed with SPSS 14.0 (Chicago, USA). A P value of less than 0.05 was considered to be statistically significant. Results Identification of a gene differentially expressed in the colorectal tumor and the adjacent normal tissue using DD-PCR The modified DD-PCR method was used to identify genes uniquely and/or highly expressed in human CRC tissues by comparing with those in adjacent normal tissues.
One cDNA band (size ranging between 700 bp and 800 bp) was found to be highly expressed in colorectal cancer tissues while barely expressed in matched adjacent normal tissues. The identified band was recovered, re-amplified, subcloned and sequenced. BLAST analysis of this nucleotide sequence revealed 99% homology to the gene DHX32 in the GenBank database. Confirmation of DHX32 differently expressed in colorectal tumors (CT) and their adjacent normal tissues (ANT) by real-time PCR We compared both positive rate and gene expression level in order to confirm the difference of DHX32 gene expression between colorectal tumors and their adjacent normal tissues. We found that the positive rate of DHX32 gene expression was
significantly higher in the colorectal tumors (76.5%) than that in the adjacent normal tissues (26.4%) (Table 2). Consistent with the higher positive rate of DHX32 gene expression in the colorectal tumors, its gene expression level was also significantly higher than that in the adjacent normal tissues (Figure 1). In addition, the distribution of the patients with decreased, constant or increased gene expression of DHX32 was also analyzed. Liothyronine Sodium Whether it was decreased, constant or increased expression for each patient was arbitrarily classified according to the gene expression ratios between the tumor tissue and its adjacent normal tissue in each patient as described in the literature [16]: decreased expression (<0.8), constant expression (0.8~1.2), increased expression (>1.2). Tumor tissues from 58.8% of the patients displayed increased gene expression of DHX32. Those from 29.4% of the patients did not change and those from 11.8% of the patients even showed decreased gene expression of DHX32 (Table 3).