Project description:Proteomic Analysis. The proteomic expression of CGMCC 6315 under different nutrient concentration conditions was investigated by isobaric tags for relative and absolute quantitation (iTRAQ)-based quantitative proteomics. The CGMCC 6315 was cultured in LB broth and 1/15LB broth as described above, after which the strains were collected by centrifugation at 10,000× g for 10 min at 4°C. Protein extraction, digestion, iTRAQ labeling and peptide fractionation were performed using the protocol described by Jin et al. 29. Protein identification was conducted using a LC-20AD nano-HPLC instrument (Shimadzu, Kyoto, Japan) equipped with a Q EXACTIVE tandem mass spectrometer (Thermo Fisher Scientific, San Jose, CA, USA) for data-dependent acquisition detection by nano-electrospray ionization. The raw MS/MS data were converted into MGF format by the thermo scientific tool Proteome Discoverer, and the exported MGF files were searched using Mascot (version 2.3.02) against the selected database containing 7546 CGMCC 6315 coding genes. The IQuant software was used for quantitative analysis of the labeled peptides with isobaric tags. Fold changes of >1.7 with p-values <0.05 were used as a cut off for differentially regulated proteins.
Project description:Arthrobacter sp. CGMCC 3584 are able to produce high yields of extracellular cyclic adenosine monophosphate (cAMP), which plays a vital role in the field of treatment of disease and animal food, during aerobic fermentation. DNA array-based transcriptional analysis of Arthrobacter cells was conducted to elucidate the higher productivity of cAMP under high oxygen supply. Results showed that 14.1% and 19.3% of the whole genome genes were up-regulated and down-regulated notably, respectively. The largest group with altered transcriptional levels belonged to the group involved in carbohydrate transport and metabolism. Other large functional groups of differentially expressed genes changed significantly included amino acid transport and metabolism, inorganic ion transport and metabolism and transcription. Overall design: The agitations were set as 150 r/min and 350 r/min to construct low oxygen supply and high oxygen supply conditions, respectively, a 5-L fermenter. Samples (three biological replicates) were respectively collected at 12h, 24h, 36h and 48h under two different oxygen supply conditions for transcriptional analysis. Equal samples of each of the independent biological replicates were mixed together for DNA microarray analysis. Genes expression level ratios (high oxygen supply/low oxygen supply) ≥ 2 or ≤ 0.5 were considered to be significantly regulated.