Flot (2007) Champuru 1.0: a computer software for unraveling mixtures of two DNA sequences of unequal lengths.Molecular Ecology Notes7 (6), 974-977 [link]
Champuru makes it possible to determine the haplotypes of heterozygous individuals without cloning, simply by analyzing the patterns of double peaks in the forward and reverse chromatograms.
Forward sequence:
The forward sequence (IUPAC one-letter code) of the heterozygous individual.
Reverse sequence:
The reverse sequence (IUPAC one-letter code) of the heterozygous individual.
Score calculation:
Which score calculation method to use. Currently the following methods are implemented:
Paper
The score correction method described in the Champuru 1.0 paper.
Ambiguity correction
A modification of the score correction method described in the Champuru 1.0 paper.
The score will get corrected for the fact that ambiguous characters (e.g. W) can match multiple other characters (e.g. A, T).
Preliminary results suggests that this score calculation method works better when the reconstructed consensus sequences contain a lot of ambiguities.
However this score correction method seems to work less good on short input sequences.
Longest Length
Take the longest number of consecutive matching nucleotides as score.
To get a quick feeling of how Champuru works, use the following data:
Forward sequence: ATSYKRMKY
Reverse sequence: WWKCTGAST
This example may be used in order to explain how Champuru can resolve ambiguities in the consensus sequences (Step 3):
Forward sequence: CTRAATTCAAATCACACTCGCGAAAWYMWKRAA
Reverse sequence: YWRAWTYMAAWYMMMMYYSSSRAAATCATGAA
A more complex example with lots of ambiguities in the consensus sequence:
Forward sequence: AAATSSYKRWTYMMMMMRSSRMSGSCCYWRSMWWMCCSRRGGRWYSGRARR
Reverse sequence: MRMTGMTKMWYMMMRCRRCGRCSSYMSYAKMMYMSMSGRRKSRKMRGRRKA
And here is a 'real-life' example (original chromatograms: Forward and Reverse):
Forward sequence: ATCGTGGGCGGCCGGCCCTGCCACAGACGGGGTTGTCACCCTCTGCGACGTGCCGTTCCAAGCAACTTAGGCAGGGCCCCTCCGCCTAGAAAGTGCTTCTCGCAACTACAACTCGCCGATGCAGGCATCGGAGATTTCAAATTTGAGCTCTTCCCGCTTCACTCGCCGTTACTGGGGGAATCCTTGTTAGTTTCTTTTCCTCCGCTTATTAATATGCTTAAATTCAGCGGGTAGCCTTGCCTGATCTGAGGTCTGGAAGGCGATTCCTTTTTTCCTTTGAGATGCCGCCACCGCTACCCGGCGGCAGCAGAAAAAAGAATCGAATGGAGAAAGATTTGTTCCGTCAAAGCGATAGAGCCGTGGCCGTTTGGGGTACATTGTTCTATGATCCCCGCGCGACACCGGATGTCGCTTGGCGGATCTTTCTCCCTGAATTTCAAGGGACGCGGTAAACCGACCGGTCGGGCCGAGCAGCACCAGGGCTGGCTAGCTAGCGCACGACCGGTCATCTCGACCGCGACCCTCAACGCCGCACGAACCCGTTCACGGCGGGCGCGYSYCSSSSCCMYMYSCKMYASASRSGGRSMMSRSGSRSRCGCGCRCRCGSRKWYKCRCRMKRKRKRTKTKWRWAKASACWCWSASASASAYRYKCYYSKGRGARMMCMMRAGMGCSMYWTKYGYKYWMARAKWYKMKRWKWYWCWSWRWWYTCYGCWWYTCACWCTWYWTMTCRSMMTCKMKCTGA
Reverse sequence: GTGCRMTMTCAACACMCSASTCTCGMRACRCATMKYGKGSGSSSSSSSCYSYSMCASASRSGGKKKTSWCMCYCTSYGMSRYGYSSYKYYMMRMRMMWYWKRSRSRGSSCCYCYSCSYMKARARWGYKYYTCKCRMMWMYAMMWCKCSSMKRYRSRSRYMKSRGAKWTYWMAWWTKWGMKCTYYYCSCKYYWCWCKCSSYKWYWSKGGGRRWMYYYKTKWKWKTYTYTTYYYCYSCKYWTWWWWATRYKYWWAWWYWSMGSGKRKMSYYKYSYSWKMTSWGRKSTSKRRRRSGMKWYYYYTTTTYYYYTKWGAKRYSSCSMCMSCKMYMCSSSGSSRSMRSARAAAARARWMKMRWRKRGARARAKWTKTKYYSYSWMARMGMKAKAGMSSYGKSSSYKTKKGGKRYAYWKTKYTMTRWKMYCCSCGCGMSACMSSRKRTSKCKYKKSGSRKMTYTYTCYCYSWRWWTYWMRRGRSRCGSKRWAMMSMSMSSKSKSGSSSMGMRSMRCMMSRGSKSKSKMKMKMKMGCRCRMSMSSKSWYMTCKMSMSCGMSMCYCWMMRCSSCRCRMRMMCSYKYWCRSSGSGSGCGCGTCCCGGCCCCATCCGCTACAGACGGGGACCAGGCGGACGCGCGCACGCGGATTCGCACGATGGATGTTTGAATAGACACTCAGACAGACATGCTCCTGGGAGAACCCAAGAGCGCCATTTGCGTTCAAAGATTCGATGATTCACTGAAT