METHODS AND COMPOSITIONS FOR COMBINATORIAL BARCODING
The present disclosure provide compositions, methods and kits for generating a set of combinatorial barcodes, and uses thereof for barcoding samples such as single cells or genomic DNA fragments. Some embodiments disclosed herein provide compositions comprising a set of component barcodes for producing a set of combinatorial barcodes. The set of component barcodes can comprise, for example, n×m unique component barcodes, wherein n and m are integers, each of the component barcodes comprises: one of n unique barcode subunit sequences; and one or two linker sequences or the complements thereof, wherein the component barcodes are configured to connect to each other through the one or two linker sequences or the complements thereof to produce a set of combinatorial barcodes.
1 . A method of barcoding a plurality of partitions comprising:
introducing m component barcodes in each of the plurality of partitions, wherein the m component barcodes are selected from a set of n×m different component barcodes, wherein n and m are positive integers, each of the component barcodes comprises:
one of n unique barcode subunit sequences; and
one or two linker sequences or the complements thereof; and
connecting the m component barcodes to generate a combinatorial barcode,
whereby each of the plurality of partitions is associated with a unique combinatorial barcode comprising the m component barcodes.
2 . The method of claim 1 , wherein each of the plurality of the partitions comprises a sample.
3 . The method of claim 2 , wherein the sample is a DNA sample or a RNA sample.
4 . The method of claim 2 , wherein the sample is a single cell.
5 . The method of claim 1 , wherein the total number of different linker sequences is m−1 or m−2.
6 . The method of claim 2 , comprising hybridizing one or two of the m component barcodes to a target of the sample.
7 . The method of claim 6 , comprising extending the one or two of the m component barcodes hybridized to the DNA target to produce an extension product comprising the combinatorial barcode.
8 . The method of claim 7 , comprising amplifying the extension product.
9 . The method of claim 8 , comprising pooling the amplification products from the plurality of partitions.
10 . The method of claim 9 , comprising sequencing the pooled amplification products.
11 . The method of claim 10 , comprising assembling the sequences using the combinatorial barcodes.
12 . The method of claim 11 , comprising generating haplotypes using the assembled sequences.
13 . The method of claim 1 , wherein the component barcodes are introduced in each of the plurality of partitions using an inkjet printer.
14 . A method of single cell sequencing comprising:
depositing a first oligonucleotide comprising a first barcode selected from a first set of 40 unique barcodes and a first primer into a plurality of partitions comprising a single cell;
depositing a second oligonucleotide comprising a second barcode selected from a second set of 40 unique barcodes and a second primer into the plurality of partitions comprising the single cell;
contacting the first oligonucleotide with a target from the single cell;
extending the first oligonucleotide to generate a first strand comprising the first barcode;
contacting the second oligonucleotide with the first strand; and
extending the second oligonucleotide to generate a second strand comprising the first barcode and the second barcode,
whereby the target from the single cell is labeled with the first barcode and the second barcode.
15 . The method of claim 14 , comprising amplifying the second strand to produce amplification products.
16 . The method of claim 15 , comprising pooling the amplification products from the plurality of partitions.
17 . The method of claim 16 , comprising sequencing the pooled amplification products.
18 . The method of claim 16 , comprising assembling the sequences from a single cell using the first barcode and the second barcode.
19 . The method of claim 14 , wherein the target is an mRNA or a DNA.
20 . The method of claim 14 , wherein the first primer is an oligo-dT primer.