Development and evaluation of 50-mer oligonucleotide arrays for detecting microbial populations in Acid Mine Drainages and bioleaching systems.
To effectively monitor microbial populations in acidic environments and bioleaching systems, a comprehensive 50-mer-based oligonucleotide microarray was developed based on most of the known genes associated with the acidophiles. This array contained 1,072 probes in which there were 571 related to 16S rRNA and 501 related to functional genes. The functional genes in the microarray were involved in carbon metabolism (158), nitrogen metabolism (72), sulfur metabolism (39), iron metabolism (68), DNA replication and repair (97), metal-resistance (27), membrane-relate gene (16), transposon (13) and IST sequence (11). Based on the results of microarray hybridizations, specificity tests with representative pure cultures indicated that the designed probes on the arrays appeared to be specific to their corresponding target genes. The detection limit was 5 ng of genomic DNA in the absence of background DNA. Strong linear relationships between the signal intensity and the target DNA were observed (r(2) approximately 0.98). Application of this type of the microarray to analyze the acidic environments and bioleaching systems demonstrated that the developed microarray appeared to be useful for profiling differences in microbial community structures of acidic environments and bioleaching systems. Our results indicate that this technology has potential as a specific, sensitive, and quantitative tool in revealing a comprehensive picture of the compositions of genes related with acidophilic microorganism and the microbial community in acidic environments and bioleaching systems, although more work is needed to improve.
Yin
Huaqun
H
School of Minerals Processing and Bioengineering, Central South University-Changsha, Hunan, 410083 China.
Cao
Linhui
L
Qiu
Guanzhou
G
Wang
Dianzuo
D
Kellogg
Laurie
L
Zhou
Jizhong
J
Dai
Zhimin
Z
Liu
Xueduan
X
D000143:Acids / Q000378:metabolism; D001419:Bacteria / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D044822:Biodiversity; D002462:Cell Membrane / Q000235:genetics; D016000:Cluster Analysis; D004260:DNA Repair / Q000235:genetics; D004261:DNA Replication / Q000235:genetics; D004251:DNA Transposable Elements / Q000235:genetics; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D004275:DNA, Ribosomal / Q000032:analysis / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D004351:Drug Resistance / Q000235:genetics; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D053858:Metabolic Networks and Pathways / Q000235:genetics; D008906:Mining; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012680:Sensitivity and Specificity; D014871:Water Microbiology
D000143:Acids; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D021903:DNA, Ribosomal Spacer; D012336:RNA, Ribosomal, 16S