Method of determining clonotypes and clonotype profiles

Abstract

The invention is directed to methods for determining clonotypes and clonotype profiles in assays for analyzing immune repertoires by high throughput nucleic acid sequencing of somatically recombined immune molecules. In one aspect, the invention comprises generating a clonotype profile from an individual by generating sequence reads from a sample of recombined immune molecules; forming from the sequence reads a sequence tree representing candidate clonotypes each having a frequency; coalescing with a highest frequency candidate clonotype any lesser frequency candidate clonotypes whenever such lesser frequency is below a predetermined value and whenever a sequence difference therebetween is below a predetermined value to form a clonotype. After such coalescence, the candidate clonotypes is removed from the sequence tree and the process is repeated. This approach permits rapid and efficient differentiation of candidate clonotypes with genuine sequence differences from those with experimental or measurement errors, such as sequencing errors.

Description

[0001]This application is a continuation-in-part of copending U.S. application Ser. No. 13 / 100,389 filed 4 May 2011, which is a continuation-in-part of copending U.S. application Ser. No. 12 / 615,263 filed 9 Nov. 2009, and this application claims benefit of U.S. provisional applications 61 / 446,822 filed 25 Feb. 2011 and 61 / 455,743 filed 25 Oct. 2010, the foregoing applications being incorporated by reference in their entireties.BACKGROUND OF THE INVENTION[0002]Analysis of biological or medical samples often requires the determination of nucleic acid sequences of large and complex populations of DNA and / or RNA, e.g. Gloor et al, PLoS ONE 5(10): el 5406 (2010); Petrosino et al, Clinical Chemistry, 55(5): 856-866 (2009); Arstila et al, Science, 286: 958-961 (1999). In particular, profiles of nucleic acids encoding immune molecules, such as T cell or B cell receptors, or their components, contain a wealth of information on the state of health or disease of an organism, so that the use of...

AI Technical Summary

Benefits of technology

[0009]The present invention provides methods for determining clonotypes and clonotype profiles from large sets of sequence data obtained by high throughput sequencing Of somatically recombined immune molecules. In one aspect, the invention implements the above methods by using one or more sequence trees for efficiently carrying out sequence comparisons necessary for distinguishing candidate clonotypes with genuine sequence differences from those with experimental or measurement errors, such as sequencing errors.

Problems solved by technology

However, the efficient determination of clonotypes and clonotype profiles from sequence data poses challenges because of the size populations to be analyzed, the limited predictability of natural variability in the sequences extracted from samples, and noise introduced into the data by a host of sample preparation and measurement steps, e.g. Warren et al, Genome Research, 21(5): 790-797 (2011).

Images