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Multiplexing Unique Dual Index Primers

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Cat.#
30075S
30075L
Index type
unique dual indexes
unique dual indexes
Reactions
96
384

The Multiplexing Unique Dual Index Primers contains primer mix for multiplexing samples for Next Generation Sequencing (NGS) on the illumina platform. Multiplexing of NGS samples will reduce sequencing costs by pooling multiple NGS libraries into a single flow cell lane.

We have developed a Four-Base Difference Index System. The system allows us to make indexes that have at least 4 bases different from each other in the 8 bases index length. Our unique dual indexing primers remove sequencing errors such as index hopping, index cross-contamination, mis-assignment of reads, amplification errors, and de-multiplexing errors. The primer set includes 96 pre-mixed unique pairs of i5 and i7 index primers in a 96-well plate (index information can be downloaded Here).

Features

  • Improves sample-identification specificity with 4-Base Difference Index System
  • Each index has at least 4 bases different from other indexes
  • 4-base difference greatly increases specificity compared to the other vendors’ 3-base difference.
  • Increases multiplexing capacity
  • 96 pre-mixed unique pairs of i5 and i7 index primers
  • Minimizes sequencing errors such as:
  • Index hopping
  • Index cross-contamination
  • Mis-assignment of reads
  • Amplification errors
  • De-multiplexing errors

Even distribution of 96 samples using dual index primers. 96 libraries were made using the BioDynami NGS DNA Library Prep Kit (Cat. # 30009) and the BioDynami Multiplexing Unique Dual Index Primers (Cat. # 30075). Libraries were pooled at equal concentration and sequenced on the illumina HiSeq 4000. The numbers of reads from 96 libraries were analyzed.

Identification of index hopping by unique dual index. 96 libraries were made using the BioDynami NGS DNA Library Prep Kit (Cat. # 30009) and the BioDynami Multiplexing Unique Dual Index Primers (Cat. # 30075). Libraries were pooled at equal concentration and sequenced on the illumina HiSeq 4000. The numbers of reads from all libraries were demultiplexed and analyzed.

  • Demultiplexed: correct i5 and i7 index combination
  • Identified index hopping: correct barcodes, but NOT correct i5 and i7 index combination
  • Other: not fitting into the above categories

Sequence of the final library with index locations:

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