Systems and methods for whole genome amplification
Provided herein are systems and methods for whole genome amplification and sequencing. In particular, provided herein are systems and methods for detection of nucleic acid variants (e.g., rare variants) in limited samples.
1. A method of whole gene amplification (WGA), the method comprising:
a) combining:
a sample of genomic DNA template;
a phi29 polymerase;
nucleic acid primers; and
dNTPs;
into a reaction mixture under conditions sufficient for genomic DNA template-directed extension of the nucleic acid primers; and
b) amplifying the extended nucleic acid primers in the reaction mixture with a thermostable DNA polymerase to generate amplified whole genome DNA,
wherein said combining and amplifying are performed in a single vessel.
2. The method of claim 1 , wherein said sample of genomic DNA is a single cell sample.
3. The method of claim 1 , wherein said sample of genomic DNA is a plurality of cells.
4. The method of claim 1 , further comprising the step of detecting one or more nucleic acid variants in said amplified whole genome DNA.
5. The method of claim 1 , wherein said method further comprises the step of performing a nucleic acid sequencing assay on said amplified whole genome DNA.
6. The method of claim 1 , wherein said amplification is conducted in a volume of 35 to 500 nL.
7. The method of claim 1 , wherein at least a subset of said primers are specific for a nucleic acid target of interest.
8. The method of claim 1 , wherein at least a subset of said primers comprises a sample barcode, molecular barcode, label, or tag sequence.
9. The method of claim 1 , wherein said vessel is a well of a microfluidic chip comprising a plurality of wells.
10. The method of claim 9 , wherein each of said wells comprises a distinct nucleic acid primer.