title: Agilent-017159 Genotypic custom 60-mer oligonucleotide multibacterial microarray
status: Public on Mar 01 2011
contact.city: Bangalore
contact.first: Genotypic
contact.last: technology
contact.email: sudha.rao@genotypic.co.in
contact.state: Karnataka
contact.address: 259, Apoorva 4th cross,80 feet Road,RMV 2ND STAGE
contact.country: India
contact.institute: Genotypic Technology
organism: synthetic construct
description: We have developed a 60-mer oligonucleotide multibacterial microarray for detection and expression profiling of biodegradative genes and bacterial diversity (16S rRNA gene) in different habitats contaminated with varieties of hazardous chemicals. The genes selected were involved in biodegradation and biotransformation of various groups of compounds viz. nitroaromatic compounds (148 genes), chloroaromatic compounds (75 genes), monoaromatic compounds (373 genes), polyaromatic hydrocarbons (174 genes), pesticides/ herbicides (34 genes), alkanes/aliphatics (185 genes) and heavy metals (68 genes), which covered a total number of 133 chemicals. The efficiency (specificity, detection sensitivity) of the developed array was evaluated using the labeled genomic DNA of pure bacterial strains, Escherichia coli DH5α and Sphingomonas sp. strain NM-05 (involved in the biodegradation of γ-hexachlorohexane isolated from IPL, Lucknow) at different concentrations of 300ng, 500ng, 800ng, 1000ng and 1250ng. The developed array was applied for profiling using the total soil DNA in five contaminated habitats of north India, viz. chloroaromatic chemicals contaminated site (India Pesticide Limited, Chinhat, Lucknow), a river sediments (Gomti river sediment, Lucknow), heavy metal industry dump site (Jajmau industrial area Kanpur), a effluent treatment plant (CETP along Ganges river near Kanpur), and an oil refinery (Mathura oil refinery). Hybridization of 16S rRNA probes revealed the presence of bacteria similar to well characterized genera involved in biodegradation of pollutants. Genes involved in complete degradation pathways for hexachlorocyclohexane (lin), 1,2,4-trichlorobenzene (tcb), naphthalene (nah), phenol (mph),  biphenyl (bph), benzene (ben), toluene (tbm), xylene (xyl), phthalate (pht), Salicylate (sal) and resistance to mercury (mer) were detected with highest intensity. The most abundant genes belonged to hydroxylases, monooxygenases and dehydrogenases which were present in all the five samples. Many compound specific genes which initiate the degradation pathway were also detected.  Thus, the array developed and validated here may be useful in assessing the biodegradative potential and composition of environmentally useful bacteria in hazardous ecosystems.
manufacture_protocol: Refer to www.opengenomics.com