Bacteriophage lambda display of complex cDNA libraries: a new approach to functional genomics.
We describe the construction and characterization of two lambda surface displayed cDNA expression libraries derived from human brain and mouse embryo. cDNA inserts were obtained by tagged random-priming elongation of commercially available cDNA libraries and cloned into a novel lambda vector at the 3' end of the D capsid protein gene, which produced highly complex repertoires (1x10(8) and 2x10(7) phage). These libraries were affinity selected with a monoclonal antibody against the neural specific factor GAP-43 and with polyclonal antibodies that recognize the EMX1 and EMX2 homeoproteins. In both cases rapid identification of specific clones was achieved, which demonstrates the great potential of the lambda display system for generating affinity selectable cDNA libraries from complex genomes.
Santi
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Istituto di Ricerche di Biologia Molecolare P. Angeletti, Via Pontina Km 30.600;, Roma, 00040 Pomezia, Italy.
Capone
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Mennuni
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C
Lahm
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Tramontano
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Luzzago
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Nicosia
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D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000915:Antibody Affinity; D010582:Bacteriophage lambda / Q000235:genetics*; D001921:Brain / Q000276:immunology / Q000378:metabolism; D002213:Capsid / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary / Q000235:genetics*; D004622:Embryo, Mammalian / Q000276:immunology / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D019922:GAP-43 Protein / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D005822:Genetic Vectors / Q000235:genetics; D016678:Genome; D018398:Homeodomain Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D019151:Peptide Library; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D014157:Transcription Factors; D000099260:Empty Spiracles Homeobox Proteins
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D000939:Epitopes; D019922:GAP-43 Protein; D018398:Homeodomain Proteins; D007106:Immune Sera; D010446:Peptide Fragments; D019151:Peptide Library; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D000099260:Empty Spiracles Homeobox Proteins