title: Palindromic sequence impedes sequencing-by-ligation mechanism.
abstract: 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.
authors.affiliation: Genomics Research Center, Academia Sinica, 128 Academia Rd,, Sec, 2, Nankang, Taipei 115, Taiwan.
authors.forename: Yu-Feng
authors.initials: YF
authors.lastname: Huang
authors.forename: Sheng-Chung
authors.initials: SC
authors.lastname: Chen
authors.forename: Yih-Shien
authors.initials: YS
authors.lastname: Chiang
authors.forename: Tzu-Han
authors.initials: TH
authors.lastname: Chen
authors.forename: Kuo-Ping
authors.initials: KP
authors.lastname: Chiu
mesh_terms: D001483:Base Sequence; D003001:Cloning, Molecular; D004277:DNA, Single-Stranded / Q000235:genetics; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D016680:Genome, Bacterial / Q000235:genetics; D055029:Inverted Repeat Sequences; D008969:Molecular Sequence Data; D017421:Sequence Analysis / Q000379:methods*
chemical_list: D004277:DNA, Single-Stranded; C049760:endodeoxyribonuclease MmeI; D015252:Deoxyribonucleases, Type II Site-Specific