Open Access Highly Accessed Research article

The application of methylation specific electrophoresis (MSE) to DNA methylation analysis of the 5' CpG island of mucin in cancer cells

Seiya Yokoyama1, Sho Kitamoto1, Norishige Yamada1, Izumi Houjou1, Tamotsu Sugai2, Shin-ichi Nakamura3, Yoshifumi Arisaka4, Kyoichi Takaori5, Michiyo Higashi1 and Suguru Yonezawa1*

Author Affiliations

1 Department of Human Pathology, Field of Oncology, Kagoshima University Graduate School of Medical and Dental Sciences, 8-35-1 Sakuragaoka, Kagoshima 890-8544, Japan

2 Division of Pathology, Central Clinical Laboratory, School of Medicine, Iwate Medical University, Morioka, Japan

3 DPR Co., LTD. 4-10-53, Mitake, Morioka 020-0122, Japan

4 Second Department of Internal Medicine, Osaka Medical College, 2-7 Daigaku-machi, Takatsuki, Osaka 569-8686, Japan

5 Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Kyoto University Graduate School of Medicine, 54 Kawara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan

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BMC Cancer 2012, 12:67  doi:10.1186/1471-2407-12-67

Published: 14 February 2012

Additional files

Additional file 1:

Table S1. Synthetic oligonucleotides used in RT-PCR.

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Additional file 2:

Table S2. MassARRAY analysis of MUC1 promoter region at Caco2 and T-47D.

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Additional file 3:

Table S3. Sequence of MUC1 promoter region, and bisulfite-sequence of Sss I treated DNA of Caco2 and PCR amplicon of T-47D.

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Additional file 4:

Figure S1. Preparation of fully methylated and unmethylated controls. The methyltransferase (Sss I) treatment of 1 μg DNA was performed at 37°C for 4 h. The fully unmethylated control was construct by PCR with the following primers (forward primer 1: 5'-CATTATCCAGCCCTCTTATTTCTC-3' and reverse primer 2: 5'-ACTTCTCTACAGGACATTTGCTTG-3') using 20 ng of DNA as template. Then, these DNA samples were applied to bisulfite treatment.

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Additional file 5:

Figure S2. Analysis of the methylation status of DNA extracted from PDAC patient. Five concentration samples were prepared by a ten-fold serial dilution using initial bisulfite treated DNA sample (20 ng/μl). D.W.: using distilled water; N.T.: using non-bisulfite treated DNA.

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Additional file 6:

Figure S3. MSE analysis of MUC2 and MUC4 promoter DNA mathylation status using human colonic normal and neoplastic crypts. N:normal tissue. T: tumor tissue. All isolated crypt samples showed high expression levels of MUC2 and MUC4 mRNA and protein.

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Additional file 7:

Figure S4. MSE analysis of MUC2 and MUC4 promoter DNA methylation status using human fluid samples. Pancreatic juice samples were collected from 2 patients with PDAC and 5 patients with IPMN. The level of methylation in PDAC was significantly lower than that in IPMN.

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