Quantitative Analysis of Bacterial Biofilm and Virulence Genes

Bacterial biofilm-related genes (icaA, icaC, icaR, SigB), autolysis-related gene (SarA), cell wall synthesis-related genes (MurE, MurC, SaeR), resistance-related gene (MecA), and bacterial virulence-related genes (ssaA, Empb) were selected for quantitative analysis by RT-PCR. MRSA bacteria was centrifuged after incubation in LB broth medium for 12 h at 37 °C and resuspended in TRIzol regent containing 100 µg/mL lysostaphin. Total RNA was extracted using RNasy mini kit (Qiagen, Düsseldorf, Germany) and then converted to cDNA using PrimeScript RT reagent kit (TaKaRa) according to the manufacturer’s instructions. The RT-PCR reaction using cDNA templates were performed with TaKaRa TB Green™ Premix Ex Taq™ II (Tli RNaseH Plus; RR820A) on QuantStudio1 applied biosystems. The cycle threshold values of all tested genes were normalized using GAPDH as reference gene, gene expressions were calculated by 2⁽⁻^ΔΔ^CT) method [67 (link)]. The primers [68 (link)] used are listed in Table S1.

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Zhang W., Margarita G.E., Wu D., Yuan W., Yan S., Qi S., Xue X., Wang K, & Wu L. (2022). Antibacterial Activity of Chinese Red Propolis against Staphylococcus aureus and MRSA. Molecules, 27(5), 1693.

Publication 2022

RNasy mini kit PrimeScript RT reagent kit TB Green™ Premix Ex Taq™ II

Autolysis Bacteria Bacterial genes Biofilm Cdna Cell wall Gapdh Gene Gene expressions Icar Lysostaphin Meca Mrsa Primers Rt pcr Synthesis Synthesis genes Trizol Virulence

Corresponding Organization : Chinese Academy of Agricultural Sciences

Other organizations : Leeds Beckett University, Shanxi Agricultural University, Liaocheng University

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Variable analysis

dependent variables

Gene expression levels of the selected genes

control variables

GAPDH as reference gene

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