Isolation of expressed genes in microorganisms

Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Preparing compound containing saccharide radical

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435 6, 435 911, 435 9121, 536 231, 536 232, 536 237, 536 2433, C12P 1934, C12Q 168, C07H 2104

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06090593&

ABSTRACT:
The differential display (DD) technique, widely used previously for eukaryotic gene discovery, is optimized to detect differential mRNA transcription (an expressed gene) from both pure culture and soil derived bacterial RNA. A model system using toluene induction of TodC1 in Pseudomonas putida F1 is used to optimize the procedure. Once optimized, an arbitrary primer for the RT step in conjunction with the same arbitrary primer and a Shine-Dalgarno (SD) primer for the PCR reaction is used to detect the expressed gene. The invention thus provides a method for discovery and acquisition of novel genes from environmental microbial communities that avoids the traditional steps and inherent bias due to the culturing of environmental isolates.

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