Selective elution of immobilized multiplexed primer extension products
View Patent ↗The present invention provides methods and kits for the sequential elution of families of extension products from one or more substrates. The methods and kits can be used, for example, to separate families of primer extension products generated by multiplex sequencing reactions or by other multiplex primer extension reactions.
1. A method of isolating a plurality of families of primer extension products from a capture substrate comprising the steps of:
(a) contacting a plurality of captured families of primer extension products with a first specific elution compound thereby selectively eluting a first family of primer extension products captured on the capture substrate, wherein the first specific elution compound is a polynucleotide; and
(b) contacting the remainder of the plurality of captured families of primer extension products with a second specific elution compound thereby selectively eluting a second family of primer extension products captured on the capture substrate.
2. The method of claim 1 wherein the second specific elution compound is a polynucleotide.
3. The method of claim 2 wherein the families of primer extension products are polynucleotide sequencing ladders.
4. The method of claim 2 wherein the families of primer extension products are polynucleotide amplification products.
5. The method of claim 2 wherein the capture substrate is a continuous solid support.
6. The method of claim 2 wherein the capture substrate comprises a plurality of solid supports.
7. The method of claim 6 wherein the capture substrate comprises a plurality of beads.
8. The method of claim 6 wherein each solid support comprises a plurality of different capture compounds.
9. A method of isolating a plurality of families of primer extension products comprising the steps of:
(a) contacting a plurality of families of primer extension products with a capture substrate, wherein
(i) a first family of primer extension products comprises a first prey moiety,
(ii) a second family of primer extension products comprises a second prey moiety, and
(iii) said capture substrate comprises a first capture compound that is capable of specifically binding the first prey moiety, and a second capture compound that is capable of specifically binding the second prey moiety,
under conditions wherein the first family of primer extension products is capable of selectively binding the first capture compound and the second family of primer extension products is capable of selectively binding the second capture compound,
(b) selectively eluting a first family of primer extension products and a second family of primer extension products from the capture substrate according to claim 2 .
10. The method of claim 9 wherein the first and second prey moieties are polynucleotide.
11. The method of claim 9 wherein the first and second capture compounds are polynucleotide.
12. A method for isolating a plurality of families of primer extension products comprising the steps of:
(a) generating a plurality of families of primer extension products from a plurality of capturable primers and a polynucleotide template, and
(b) isolating the plurality of families of primer extension products according to claim 9 .
13. A method for isolating a plurality of families of primer extension products comprising the steps of:
(a) generating a plurality of families of primer extension products from a plurality of capturable primers and a plurality of polynucleotide templates, and
(b) separating the plurality of families of primer extension products according to claim 9 .