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Published on: April 11, 2016
Primer-walking sequencing identifies predominantly noncoding TP53 variants in canine cancers
Ramya Mathiyalagan1, Chanjoong Lee2, Yongsuk Jo2
1Department of Chemistry, Chungnam National University, Daejeon 34134, Korea.
Importance:
Tumor protein p53 (TP53) alterations are frequently reported in canine cancers but remain incompletely characterized due to differences in reference genomes and sequencing cost.
Objective:
To evaluate a cost-efficient primer-walking approach for TP53 variant screening in dogs with spontaneous tumors.
Methods:
Tumor tissue (n = 10) or blood (n = 16) was collected from 26 dogs with suspected neoplasia. Interpretable TP53 sequences spanning introns and exons 2-11, excluding long intron 1 and non-protein-coding exon 1, were obtained by primer-walking polymerase chain reaction and Sanger sequencing. Bidirectional reads were compared with the Ensembl ROS_Cfam_1.0 v114 reference sequence.
Results:
TP53 gene sequence variations were observed in 18/26 sequenced tumor cases, including one exonic frameshift insertion and 17 noncoding variants: 13 intronic variants and 4 variants in the 5' untranslated region. Recurrent substitutions and deletions occurred in the intron 8-10 region. Variants were detected in lymphomas and carcinomas, including cases from several small breeds, but no significant association with tumor type or breed was demonstrated.
Conclusions And Relevance:
Primer-walking Sanger sequencing can screen TP53 regions beyond coding exons in canine tumor samples. Matched tumor-normal sequencing is needed to determine somatic versus germline origin and to evaluate noncoding TP53 variants as potential biomarkers or drivers.
