Patent EP-3474169-A1 (EP)
Inventor: EZPELETA JOAQUÍN
IPC: G06N3/12
Filed: 2017-10-20
Abstract: A first aspect of the present invention relates to a method of tagging a nucleic acid molecule with a predetermined ID number, the method comprising: (a) attaching to said nucleic acid molecule a nucleic acid tag to form a tagged nucleic acid molecule, wherein said nucleic acid tag comprises one or more nucleic acid tag sub-units each consisting of groups of at least two nucleotides, wherein said nucleic acid tag is attributed said ID number by performing the following steps: (i) converting each nucleotide in said nucleic acid tag sub-units into a number ranging from 0 to 3, thereby creating numerical tag sub-units, wherein the distribution and content of the nucleotides in the nucleic acid tag sub-units has been configured to allow a finite number of numerical tag sub-units, (ii) attributing to each of said numerical tag sub-units a predetermined numerical tag element, thereby creating a numerical tag, (iii) linearly decoding said numerical tag, thereby creating said ID number.  A second aspect of the present invention relates to a method for multiplex sequencing and/or demultiplexing, the method comprising the steps of: (a) multiplexing amplification of tagged nucleic acid molecules obtained according to the method of the first aspect of the present invention to generate a plurality of said tagged nucleic acid molecules, (b) pooling and parallel sequencing said plurality of tagged nucleic acid molecules, thereby generating a plurality of sequence reads, and (c) demultiplexing said plurality of sequence reads, wherein each of said sequence reads is attributed to a sample.  A third aspect of the present invention is directed to a tagged nucleic acid molecule construed according to the method of the first aspect of the present invention and its use in a method for multiplex sequencing and/or demultiplexing.  A fourth aspect of the present invention relates to an apparatus configured for multiplex sequencing demultiplexing, the apparatus comprising: tools for designing nucleic acid tags according to the method described in the first aspect of the present invention, tools for pooling and multiplexing a plurality of tagged nucleic acid sequences, a sequence demultiplexer, additional tools for data reproduction and post-sequencing analysis.  A fifth aspect of the present invention, there is provided a method of tagging a nucleic acid molecule with a predetermined ID number, the method comprising: (a) attaching to said nucleic acid molecule a nucleic acid tag to form a tagged nucleic acid molecule, wherein said nucleic acid tag comprises one or more nucleic acid tag sub-units each consisting of groups of at least two nucleotides, wherein said nucleic acid tag is obtained from said ID number by performing the following steps: (i) linearly encoding said ID number, thereby creating a numerical tag, wherein the numerical tag comprises a plurality of numerical tag elements; (ii) attributing to each of said numerical tag elements a numerical tag sub-unit, wherein the numerical tag sub-unit comprises a plurality of numbers ranging from 0 to 3; (iii) attributing to each of said numerical tag sub-units a nucleic acid tag sub-unit, thereby creating said nucleic acid tag, wherein the distribution and content of the nucleotides in the nucleic acid tag sub-units has been configured to allow a finite number of numerical tag sub-units.