TMI-seq: Tn5 Transposase Mediated Production of Complex Libraries for Short Read Sequencing

The technology, termed “TMI-seq” is a library preparation that combines molecular barcoding of individual molecules with “tagmentation” – a process by which the TN5 transposase both fragments and adds tags to DNA. In one u…

The technology, termed “TMI-seq” is a library preparation that combines molecular barcoding of individual molecules with “tagmentation” – a process by which the TN5 transposase both fragments and adds tags to DNA. In one use of the method, RNA is reverse transcribed to integrate a molecular barcode and a partial forward and reverse sequencing primer. It is then amplified by PCR, which integrates index sequences and full forward and reverse sequencing primers to yield a full length cDNA library. One aliquot of the library is tagmented with Tn5 and the forward sequencing primer, resulting in a library of barcoded overlapping fragments focused on the 5’ region of the target sequence. A second aliquot is tagmented with Tn5 and the reverse sequencing primer, resulting in a library of barcoded overlapping fragments focused on the 3’ region of the target sequence.

Abstract:

Although Next Generation Sequencing has vastly improved sequencing throughput while reducing sequencing costs, the preparation of nucleic acid libraries for sequencing has become a bottleneck. In addition, it is difficult using short-read next-generation sequencing to assemble highly variable sequences that exceed 500 base pairs such as cDNAs derived from antibody heavy chain, antibody light chain, and T cell variable regions RNA.

Website:

https://techtransfer.universityofcalifornia.edu/NCD/32761.html?utm_source=AUTMGTP&utm_medium=webpage&utm_term=ncdid_32761&utm_campaign=TechWebsites

Advantages:

  • Fast prep and accurate assembly for short read sequencers like Illumina sequencers. 
  • High quality data from immune repertoire sequences
  • Accurate TCR and antibody repertoire sequences

Potential Applications:

  • RNA à cDNA library sequencing
  • Sequencing of T Cell receptors and antibodies

Contact Information:

Name: Jeff Jackson

Email: jjackso6@ucsc.edu

Phone: (831) 459-3976