Improved transposon-based library preparation for the Ion Torrent platform.

Area of Science:

• Molecular Biology
• Next-Generation Sequencing

Background:

• Transposon-based library preparation is efficient for Illumina and Ion Torrent platforms.
• PCR-mediated adaptor incorporation can form hairpin structures and heteroduplexes, reducing Ion Torrent data output by over 3-fold.

Purpose of the Study:

• To develop a novel transposon-based protocol for barcoded library preparation for the Ion Torrent platform.
• To eliminate hairpin formation and heteroduplexes in sequencing libraries.

Main Methods:

• Developed MuPlus, a protocol integrating one adaptor by PCR and the second by ligation of a single-stranded oligonucleotide.
• Enzymatic cleavage of a complementary part on one strand of the tag facilitates single-stranded adaptor ligation.

Main Results:

• The MuPlus protocol generates libraries free of self-complementary, hairpin-forming regions.
• Libraries prepared with MuPlus are free of heteroduplexes.
• MuPlus libraries achieve sequencing efficiency comparable to ligation-based protocols on the Ion Torrent platform.

Conclusions:

• MuPlus offers an improved transposon-based method for Ion Torrent library preparation.
• The protocol effectively overcomes limitations associated with PCR-mediated adaptor incorporation.
• MuPlus enhances sequencing data output and reliability for the Ion Torrent platform.