RNA was extracted from the pseudomonas aeruginosa isolates (P1, P5, P9, P17, P18, and P29) before treating as a control and after treating with the AgNPs Aloe vera plant Extract. Total RNA was extracted using TRIzol™ Reagent and its concentration was from 80 to 210 ng/μL, the purity of total RNA samples ranged from (1.6 to1.9). mexZ and aac6 genes which are carried by the six P. aeruginosa isolates (P1, P5, P9, P17, and P18) that were used in the investigation of gene expression. These isolates were chosen because they exhibited the highest level of resistance. The influence of the AgNPs Aloe vera leaves extract on the Gene Expression of the mexZ and aac6 Genes in P. aeruginosa Isolates was the primary focus of this research. Primers designed to target the mexZ and aac6 genes in P. aeruginosa have a high degree of specificity for gene expression. P. aeruginosa isolates had their gene expression of the aac6 and mexZ genes measured using real-time PCR before and after being treated with the AgNPs Aloe vera plant extract. Reverse transcription-quantitative PCR (RT-qPCR) was carried out using SYBR green; a fluorescent dye that recognizes any double-stranded DNA including cDNA. Through a two-step RT-qPCR method in order to determine the effect of sub MICs of AgNPs Aloe vera leaves on the expression of mexZ and aac6 gene in P. aeruginosa. The first step is Synthesis cDNA from RNA by used specific primer (hexamer) for the target gene and housekeeping gene in the study, the immediately transformed of genomic RNA to cDNA after extraction accomplished to get a complete transformation without any loss or degradation of RNA in one steps of cDNA synthesis. The second step is using the quantitative PCR (RT-qPCR) device to measure the range of Ct value of this gene under study. The result of real-time quantitative PCR experiments for gene aac6 was inhibited in the gene expression after treatment with AgNps Aloe vera leaves extract. The result of real-time quantitative PCR experiments for gene mexZ was inhibited in the gene expression after treatment with AgNps Aloe vera leaves extract. To reveal the inhibition efficacy of AgNps Aloe vera leaves extract on the P.aeruginosa gene, mexZ and aac6 expression were measured. The results when the isolates were treated with AgNps Aloe vera plant extract with sub-MIC (10 mg/mL) for each sample. It was found that the presence of AgNps Aloe vera leaves extract negatively affects in mexZ and aac6 gene expression. Decreased expression in mexZ and aac6 genes causes to reduce of the efflux pump expression and production of enzyme modify which is one of the causes of antibiotic resistance. Decreased mexZ and aac6 gene expression is an important indicator that it influences an important mechanism of resistance in P. aeruginosa. The main purpose of studying gene expression in the presence and absence of AgNps Aloe vera extract is to investigate the role of these genes in efflux pump formation and enzyme modification production. It is considered one of the most important virulence genes that cause resistance to the aminoglycoside group in P. aeruginosa. Generally, numerous studies have shown that Aloe vera helps wound healing because of its anti-inflammatory, antioxidant, and immuneomodulatory activities