Measurement and monitoring of cell clonality
Methods are provided for the detection and analysis of clonality in a cell population, where parallel sequencing is applied to a nucleic acid sample obtained from the cell population, optionally a population of lymphocytes. Replicate samples are amplified, and sequenced, where identification of coincident sequences in two or more replicates is indicative of clonal expansion.
1. A method of determining the clonal expansion of B cells in response of an individual to a vaccine, the method comprising:
obtaining a cell sample comprising B cells from said individual in a short defined time period following immunization with said vaccine;
dividing said cell sample comprising B cells, or genomic DNA derived therefrom, into at least two distinct pools;
amplifying genomic DNA sequences at an immunoglobulin heavy chain locus in said at least two distinct pools with a primer set that amplifies at least 50% of the known rearrangements at the locus;
sequencing at least 10 3 reads of the amplified genomic DNA at the locus of interest from said at least two distinct pools;
comparing sequences from said amplified genomic DNA to detect the presence of sequences that are coincident in said at least two distinct pools of nucleic acid;
wherein the presence of coincident sequences is indicative of clonal expansion and responsiveness to said vaccine.
2. A method of determining the clonal expansion of B cells in response of an individual to a vaccine, the method comprising:
obtaining a cell sample comprising B cells from said individual in a short defined time period following immunization with said vaccine wherein said short defined time period is less than about 14 days;
dividing said cell sample comprising B cells, or genomic DNA derived therefrom, into at least two distinct pools;
amplifying genomic DNA sequences at an immunoglobulin heavy chain locus in said at least two distinct pools with a primer set that amplifies at least 75% of the known rearrangements at the locus;
sequencing at least 10 3 reads of the amplified genomic DNA at the locus of interest from said at least two distinct pools;
comparing sequences from said amplified genomic DNA to detect the presence of sequences that are coincident in said at least two distinct pools of nucleic acid;
wherein the presence of coincident sequences is indicative of clonal expansion and responsiveness to said vaccine.
3. The method of claim 2 in which the sequences are selected by amplification using oligonucleotide primers set forth in Table 6 with homology to specific regions of the genome.
4. The method of claim 2 , wherein the short defined time period is less than about 7 days.