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**IonTom** · 05-10-2014, 08:16 AM

The reads are actually much longer. If you open your aligned BAM/SAM in IGV and make soft clipping visible you should see it. Sometimes you even see things like 235S17M.

In a recent paper they showed that for randomly generated reads with 300 to 400 base pairs
TMAP was still aligning like 15% of these.

Comparison of mapping algorithms used in high-throughput sequencing: application to Ion Torrent data - BMC Genomics

http://www.biomedcentral.com/1471-2164/15/264/

Background The rapid evolution in high-throughput sequencing (HTS) technologies has opened up new perspectives in several research fields and led to the production of large volumes of sequence data. A fundamental step in HTS data analysis is the mapping of reads onto reference sequences. Choosing a suitable mapper for a given technology and a given application is a subtle task because of the difficulty of evaluating mapping algorithms. Results In this paper, we present a benchmark procedure to compare mapping algorithms used in HTS using both real and simulated datasets and considering four evaluation criteria: computational resource and time requirements, robustness of mapping, ability to report positions for reads in repetitive regions, and ability to retrieve true genetic variation positions. To measure robustness, we introduced a new definition for a correctly mapped read taking into account not only the expected start position of the read but also the end position and the number of indels and substitutions. We developed CuReSim, a new read simulator, that is able to generate customized benchmark data for any kind of HTS technology by adjusting parameters to the error types. CuReSim and CuReSimEval, a tool to evaluate the mapping quality of the CuReSim simulated reads, are freely available. We applied our benchmark procedure to evaluate 14 mappers in the context of whole genome sequencing of small genomes with Ion Torrent data for which such a comparison has not yet been established. Conclusions A benchmark procedure to compare HTS data mappers is introduced with a new definition for the mapping correctness as well as tools to generate simulated reads and evaluate mapping quality. The application of this procedure to Ion Torrent data from the whole genome sequencing of small genomes has allowed us to validate our benchmark procedure and demonstrate that it is helpful for selecting a mapper based on the intended application, questions to be addressed, and the technology used. This benchmark procedure can be used to evaluate existing or in-development mappers as well as to optimize parameters of a chosen mapper for any application and any sequencing platform.

If TMAP finds subsequences that align to the reference (like 17bp of 300) it will soft clip from the 5' end and report the remaining part of the read.

I don't know if is this is a bug or on purpose, but it kind of made me stop using TMAP.

**snetmcom** · 05-17-2014, 09:54 AM

Originally posted by IonTom View Post

The reads are actually much longer. If you open your aligned BAM/SAM in IGV and make soft clipping visible you should see it. Sometimes you even see things like 235S17M.

In a recent paper they showed that for randomly generated reads with 300 to 400 base pairs
TMAP was still aligning like 15% of these.

Comparison of mapping algorithms used in high-throughput sequencing: application to Ion Torrent data - BMC Genomics

http://www.biomedcentral.com/1471-2164/15/264/

Background The rapid evolution in high-throughput sequencing (HTS) technologies has opened up new perspectives in several research fields and led to the production of large volumes of sequence data. A fundamental step in HTS data analysis is the mapping of reads onto reference sequences. Choosing a suitable mapper for a given technology and a given application is a subtle task because of the difficulty of evaluating mapping algorithms. Results In this paper, we present a benchmark procedure to compare mapping algorithms used in HTS using both real and simulated datasets and considering four evaluation criteria: computational resource and time requirements, robustness of mapping, ability to report positions for reads in repetitive regions, and ability to retrieve true genetic variation positions. To measure robustness, we introduced a new definition for a correctly mapped read taking into account not only the expected start position of the read but also the end position and the number of indels and substitutions. We developed CuReSim, a new read simulator, that is able to generate customized benchmark data for any kind of HTS technology by adjusting parameters to the error types. CuReSim and CuReSimEval, a tool to evaluate the mapping quality of the CuReSim simulated reads, are freely available. We applied our benchmark procedure to evaluate 14 mappers in the context of whole genome sequencing of small genomes with Ion Torrent data for which such a comparison has not yet been established. Conclusions A benchmark procedure to compare HTS data mappers is introduced with a new definition for the mapping correctness as well as tools to generate simulated reads and evaluate mapping quality. The application of this procedure to Ion Torrent data from the whole genome sequencing of small genomes has allowed us to validate our benchmark procedure and demonstrate that it is helpful for selecting a mapper based on the intended application, questions to be addressed, and the technology used. This benchmark procedure can be used to evaluate existing or in-development mappers as well as to optimize parameters of a chosen mapper for any application and any sequencing platform.

If TMAP finds subsequences that align to the reference (like 17bp of 300) it will soft clip from the 5' end and report the remaining part of the read.

I don't know if is this is a bug or on purpose, but it kind of made me stop using TMAP.

These are short PCR products, so I doubt it's from alignment. He is likely seeing these in the histogram from the report.

There are no settings for 5' trimming. I'm guessing it's primer dimer or he means 3' trimming.

**IonTom** · 05-17-2014, 11:43 PM

Originally posted by snetmcom View Post

These are short PCR products, so I doubt it's from alignment. He is likely seeing these in the histogram from the report.

There are no settings for 5' trimming. I'm guessing it's primer dimer or he means 3' trimming.

It should also be noted that Ion Torrent produces a relatively large range of different
read lenghts i.e 30-330.

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