A cad-gfpmut3 plasmid construct in Escherichia coli for gene induction-based quantification of lysine in acid hydrolysates of feedstuffs

Abstract

To generate an inducible plasmid-borne cad-gfpmut3 transcriptional fusion and develop a method for quantification of total lysine. The cad-gfpmut3 transcriptional fusion was constructed by cloning the cad promoter (Pcad) upstream of a promotorless gfpmut3 located on a high-copy plasmid. The construct was electroporated into Escherichia coli ZK126 and the transformed strain was subsequently used to quantify lysine in feed ingredients. Lysine standard curves based on gene induction of the bacterial cells were used for estimating acid hydrolysate lysine concentrations in four feed ingredients. Except for sorghum, no substantial differences were observed when the data for lysine in soybean (2 x 49 +/- 0 x 37%), cottonseed (1 x 82 +/- 0 x 15%), and meat and bone meal (2 x 31 +/- 0 x 24%) generated by the newly developed construct were compared with previously published data. Using the cad-gfpmut3 fusion, feed derived lysine induction was measured easily and accurately, and could be a useful tool for the estimation of lysine in acid hydrolysates of feed ingredients. The described approach for lysine quantification in feed ingredients represents a cost- and time-efficient method offering rapid and accurate lysine quantification of multiple samples.