Mixed sequence reader: a program for analyzing DNA sequences with heterozygous base calling.
The direct sequencing of PCR products generates heterozygous base-calling fluorescence chromatograms that are useful for identifying single-nucleotide polymorphisms (SNPs), insertion-deletions (indels), short tandem repeats (STRs), and paralogous genes. Indels and STRs can be easily detected using the currently available Indelligent or ShiftDetector programs, which do not search reference sequences. However, the detection of other genomic variants remains a challenge due to the lack of appropriate tools for heterozygous base-calling fluorescence chromatogram data analysis. In this study, we developed a free web-based program, Mixed Sequence Reader (MSR), which can directly analyze heterozygous base-calling fluorescence chromatogram data in .abi file format using comparisons with reference sequences. The heterozygous sequences are identified as two distinct sequences and aligned with reference sequences. Our results showed that MSR may be used to (i) physically locate indel and STR sequences and determine STR copy number by searching NCBI reference sequences; (ii) predict combinations of microsatellite patterns using the Federal Bureau of Investigation Combined DNA Index System (CODIS); (iii) determine human papilloma virus (HPV) genotypes by searching current viral databases in cases of double infections; (iv) estimate the copy number of paralogous genes, such as β-defensin 4 (DEFB4) and its paralog HSPDP3.
Department of Computer Science, National Tsing Hua University, Hsin-Chu, Taiwan.
Chun-Tien
CT
Chang
Chi-Neu
CN
Tsai
Chuan Yi
CY
Tang
Chun-Houh
CH
Chen
Jang-Hau
JH
Lian
Chi-Yu
CY
Hu
Chia-Lung
CL
Tsai
Angel
A
Chao
Chyong-Huey
CH
Lai
Tzu-Hao
TH
Wang
Yun-Shien
YS
Lee
D000465:Algorithms; D020029:Base Pairing / Q000235:genetics*; D001483:Base Sequence; D004247:DNA / Q000235:genetics*; D006580:Genetic Carrier Screening / Q000379:methods*; D020407:Internet; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA / Q000379:methods*; D012984:Software
D004247:DNA