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A key limitation in using heterologous genomic or metagenomic libraries in functional genomics and genome engineering is the low expression of heterologous genes in screening hosts, such as  Escherichia coli . To overcome this limitation, here we generate  E. coli  strains capable of recognizing heterologous promoters by expressing heterologous sigma factors. Among seven sigma factors tested, RpoD from  Lactobacillus plantarum  ( Lpl ) appears to be able of initiating transcription from all sources of DNA. Using the promoter GFP-trap concept, we successfully screen several heterologous and metagenomic DNA libraries, thus enlarging the genomic space that can be functionally sampled in  E. coli . For an application, we show that screening fosmid-based  Lpl  genomic libraries in an  E. coli  strain with a chromosomally integrated  Lpl rpoD  enables the identification of  Lpl  genetic determinants imparting strong ethanol tolerance in  E. coli . Transcriptome analysis confirms increased expression of heterologous genes in the engineered strain.

Expression of heterologous sigma factors enables functional screening of metagenomic and heterologous genomic libraries