Protein evolution by hypermutation and selection in the B cell line DT40.
Genome-wide mutations and selection within a population are the basis of natural evolution. A similar process occurs during antibody affinity maturation when immunoglobulin genes are hypermutated and only those B cells which express antibodies of improved antigen-binding specificity are expanded. Protein evolution might be simulated in cell culture, if transgene-specific hypermutation can be combined with the selection of cells carrying beneficial mutations. Here, we describe the optimization of a GFP transgene in the B cell line DT40 by hypermutation and iterative fluorescence activated cell sorting. Artificial evolution in DT40 offers unique advantages and may be easily adapted to other transgenes, if the selection for desirable mutations is feasible.
Arakawa
Hiroshi
H
Institute for Molecular Radiobiology, Institute of Stem Cell Research, GSF-National Research Center for Environment and Health, Ingolstaedter Landstrasse 1, D-85764 Neuherberg-Munich, Germany.
Kudo
Hiroaki
H
Batrak
Vera
V
Caldwell
Randolph B
RB
Rieger
Michael A
MA
Ellwart
Joachim W
JW
Buerstedde
Jean-Marie
JM
D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology; D001483:Base Sequence; D002460:Cell Line; D002469:Cell Separation; D002645:Chickens / Q000276:immunology; D004252:DNA Mutational Analysis; D019020:Directed Molecular Evolution / Q000379:methods*; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000032:analysis; D018390:Gene Targeting; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics*; D007147:Immunoglobulin Light Chains / Q000235:genetics; D008969:Molecular Sequence Data; D015202:Protein Engineering / Q000379:methods*; D027041:Somatic Hypermutation, Immunoglobulin; D013050:Spectrometry, Fluorescence; D019076:Transgenes
D005456:Fluorescent Dyes; D007147:Immunoglobulin Light Chains; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins