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Palindromic sequence impedes sequencing-by-ligation mechanism
BACKGROUND: Current next-generation sequencing (NGS) platforms adopt two types of sequencing mechanisms: by synthesis or by ligation. The former is employed by 454 and Solexa systems, while the latter by SOLiD system. Although the pros and cons for each sequencing mechanism have more or less been di...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3521181/ https://www.ncbi.nlm.nih.gov/pubmed/23281822 http://dx.doi.org/10.1186/1752-0509-6-S2-S10 |
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author | Huang, Yu-Feng Chen, Sheng-Chung Chiang, Yih-Shien Chen, Tzu-Han Chiu, Kuo-Ping |
author_facet | Huang, Yu-Feng Chen, Sheng-Chung Chiang, Yih-Shien Chen, Tzu-Han Chiu, Kuo-Ping |
author_sort | Huang, Yu-Feng |
collection | PubMed |
description | BACKGROUND: Current next-generation sequencing (NGS) platforms adopt two types of sequencing mechanisms: by synthesis or by ligation. The former is employed by 454 and Solexa systems, while the latter by SOLiD system. Although the pros and cons for each sequencing mechanism have more or less been discussed in a number of occasions, the potential obstacle imposed by palindromic sequences has not yet been addressed. METHODS: To test the effect of the palindromic region on sequencing efficacy, we clonally amplified a paired-end ditag sequence composed of a 24-bp palindromic sequence flanked by a pair of tags from the E. coli genome. We used the near homogeneous fragments produced from MmeI digestion of the amplified clone to generate a sequencing library for SOLiD 5500xl sequencer. RESULTS: Results showed that, traditional ABI sequencers, which adopt sequencing-by-synthesis mechanism, were able to read through the palindromic region. However, SOLiD 5500xl was unable to do so. Instead, the palindromic region was read as miscellaneous random sequences. Moreover, readable tag sequence turned obscure ~2 bp prior to the palindromic region. CONCLUSIONS: Taken together, we demonstrate that SOLiD machines, which employ sequencing-by-ligation mechanism, are unable to read through the palindromic region. On the other hand, sequencing-by-synthesis sequencers had no difficulty in doing so. |
format | Online Article Text |
id | pubmed-3521181 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-35211812012-12-14 Palindromic sequence impedes sequencing-by-ligation mechanism Huang, Yu-Feng Chen, Sheng-Chung Chiang, Yih-Shien Chen, Tzu-Han Chiu, Kuo-Ping BMC Syst Biol Proceedings BACKGROUND: Current next-generation sequencing (NGS) platforms adopt two types of sequencing mechanisms: by synthesis or by ligation. The former is employed by 454 and Solexa systems, while the latter by SOLiD system. Although the pros and cons for each sequencing mechanism have more or less been discussed in a number of occasions, the potential obstacle imposed by palindromic sequences has not yet been addressed. METHODS: To test the effect of the palindromic region on sequencing efficacy, we clonally amplified a paired-end ditag sequence composed of a 24-bp palindromic sequence flanked by a pair of tags from the E. coli genome. We used the near homogeneous fragments produced from MmeI digestion of the amplified clone to generate a sequencing library for SOLiD 5500xl sequencer. RESULTS: Results showed that, traditional ABI sequencers, which adopt sequencing-by-synthesis mechanism, were able to read through the palindromic region. However, SOLiD 5500xl was unable to do so. Instead, the palindromic region was read as miscellaneous random sequences. Moreover, readable tag sequence turned obscure ~2 bp prior to the palindromic region. CONCLUSIONS: Taken together, we demonstrate that SOLiD machines, which employ sequencing-by-ligation mechanism, are unable to read through the palindromic region. On the other hand, sequencing-by-synthesis sequencers had no difficulty in doing so. BioMed Central 2012-12-12 /pmc/articles/PMC3521181/ /pubmed/23281822 http://dx.doi.org/10.1186/1752-0509-6-S2-S10 Text en Copyright ©2012 Huang et al.; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Proceedings Huang, Yu-Feng Chen, Sheng-Chung Chiang, Yih-Shien Chen, Tzu-Han Chiu, Kuo-Ping Palindromic sequence impedes sequencing-by-ligation mechanism |
title | Palindromic sequence impedes sequencing-by-ligation mechanism |
title_full | Palindromic sequence impedes sequencing-by-ligation mechanism |
title_fullStr | Palindromic sequence impedes sequencing-by-ligation mechanism |
title_full_unstemmed | Palindromic sequence impedes sequencing-by-ligation mechanism |
title_short | Palindromic sequence impedes sequencing-by-ligation mechanism |
title_sort | palindromic sequence impedes sequencing-by-ligation mechanism |
topic | Proceedings |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3521181/ https://www.ncbi.nlm.nih.gov/pubmed/23281822 http://dx.doi.org/10.1186/1752-0509-6-S2-S10 |
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