FLER - Follicular Lymphoma Evolution Reconstructor

Code: Peter Vasil, Technical University, Berlin
Method: Martin Wartenberg, Technical University, Munich

Materials for download:



Test data:


FLER is the implementation of the correspondent mathematic/statistic algorithm to calculate the most likely evolution of monoclonal operational taxonomic units (OTU) driven by somatic hypermutation. Thus, given a group of OTUs defined by various mutation patterns of a certain DNA-sequence (compared to an unmutated wildtype configuration (germline)), FLER uses the number of equally mutated loci among subgroups of OTUs, that share different spectra of mutations, as foundation for creation of hypothetical predecessor clones (HPCs). Among the abundance of HPCs the most probable HPCs are chosen to recapitulate evolution from the wildtype configuration to each OTU. Probability is assessed by calculation of the probability measure "pm" for each HPC. Input parameters of the pm-formula are:

\[ pm = \sqrt[(t - r + 1) \cdot i \cdot z]{\prod \limits_{k=1}^{l} P_{k}} \]

r: number of OTUs in an OTU subgroup, defining the considered HPC
i: nuber of mutations of an HPC (thus number of mutations shared by all clones of the HPC-defining subgroup)
z: number of reapeats of an HPC with the identical mutation patterns among the abundance of HPCs
t: total number of OTUs
P: relative frequency of a certain mutation among the considered population of OTUs
l: sequence length
k: counter, starting at locus 1 and stepping through the sequence until the last locus l is reached

Unlike multiple sequence alignment (MSA) algorithms, like the neighbor joining (NJ) algorithm, FLER calculates the most likely HPCs from mutation frequencies and considers the mutation number as surrogate marker for elapsed time. Thus clonal interchange among different populations of OTUs can also be assessed by doing population-specific assays first, complemented by an assay comprising all OTUs as one population. By comparing pm-values of HPCs of different assays, interchange among various populations rather than population-contained evolution can be evaluated. Then, migration directions are determined by the population affiliation of the majority of HPC-defining OTUs.


Graphical user interface (GUI):


sequence length - user can determine the sequence length (all sequences must have the identical length).
GERMLINE - user can choose the germline sequence. This sequence will be regarded as unmutated wildtype configuration.
Each clone sequence will be compared to the germline sequence to determine the clone-specific mutation pattern.
CLONES - user can choose the clones, defining the sequence population for that the evolution reconstruction will be implemented by FLER.
RUN FLER - starts the algorithm
SAVE FILE - user can choose where to save the file
RESET - sets the default for a new run of FLER

Input format / data:
Individual DNA sequences have to be saved in a plain text file (.txt), each. The sequence itself has to be written in the first line only, with no figures (lower or capital case) except for A (adenine), C (cytidine), T (thymine) , G (guanine). The name of the txt-file will be used as clone name by FLER (example: the sequence contained in file "clone1.txt" will be addressed as "clone1"). FLER accounts for single nucleotid point mutations only. Insertions, deletions, dublications or other chromosomal aberrations are not considered, and thus not to be included.

Output data:
The user can choose whether to display only the pm-directed hierarchy of HPCs, the groups of OTUs defining a certain HPC, or individual clonal metatdata like mutated loci, mutation number etc. (see "save options" in the menu bar of FLER). All output data will be saved in a file the user can determine interactively. It is recommended to use the extension <.txt>.