Figure 1

Mini-gene analysis of MLH1 , MSH2 , and MSH6 intronic mutations. COS-7 cells were transfected with wild type or mutant plasmids in duplicate. Total RNA was isolated, and RT-PCR analysis was performed. PCR products were separated by agarose gel electrophoresis and visualized by ethidium bromide staining. The sizes of the DNA marker (M) are indicated to the left. All PCR products were verified by sequencing. (a) The MLH1 c.117-34A > T mutation produced a 268-bp PCR product which corresponds to wild type exon 2 (unaltered splicing). (b) The MLH1 c.588 + 5G > A mutation produced a 266-bp band corresponding to the exclusion of exon 7. (c) The MLH1 c.677 + 3A > T mutation resulted in a 220-bp band corresponding to a transcript lacking exon 8. (d) The MLH1 c.1039-8 T > A mutation produced a 548-bp PCR product which corresponds to wild type exon 12 (unaltered splicing). (e) The MLH1 c.1732-2A > T mutation resulted in a 177-bp transcript corresponding to the exclusion of exon 16. (f) The MSH2 c.1276 + 1G > T mutation produced a 329-bp transcript by usage of a cryptic splice donor site 48 bp within exon 7. (g) The MSH2 c.1662-2A > C mutation resulted in a 177-bp product corresponding to a transcript lacking exon 11. (h) The MSH2 c.2459-18delT mutation produced a 353-bp PCR product which corresponds to wild type exon 14 (unaltered splicing). (i) The MSH6 c.3439-16C > T mutation produced a 295-bp PCR product which corresponds to wild type exon 5 (unaltered splicing).