Gene expression profiling following the overexpression or the knockdown of Ambra1 in A375 melanoma cell lines. Gene expression profiling following the overexpression or the knockdown of Ambra1 in A375 melanoma cell lines

Melanoma is the most deadly type of skin cancers and, there is a continuous need to develop early biomarkers as well as treatments to improve the survivability of patients. Ambra1 is an essential regulator in autophagy and it has been recently shown to have a role in cell proliferation. We used microarrays to analyse the transcriptomic changesi n A375 melanoma cell lines following Ambra1 differential expression Overall design: Four stably transfected A375 melanoma cell lines were utilized for this study. Overexpressions of Ambra1 and its matching control, β-galactosidase were performed by retroviral infection. While knockdown of Ambra1 was performed by lentiviral infection that expresses a shRNA construct targeted to Ambra1 and the control cell line expressed a scrambled shRNA control. Total RNA was extracted and hybridized on Affymetrix microarrays. Three biological replicates of each cell line were used for eah group.

黑色素瘤（Melanoma）是致死性最强的皮肤癌亚型，学界始终亟需开发早期生物标志物与治疗方案，以提升患者的生存率。Ambra1是自噬（autophagy）的核心调控因子，近期研究证实其在细胞增殖过程中发挥重要调控作用。本研究采用微阵列（microarray）技术，分析了Ambra1差异表达后A375黑色素瘤细胞系的转录组变化。实验设计：本研究共使用4株稳定转染的A375黑色素瘤细胞系。通过逆转录病毒感染，分别完成Ambra1及其对照β-半乳糖苷酶（β-galactosidase）的过表达；通过携带靶向Ambra1的短发夹RNA（shRNA）的慢病毒感染，实现Ambra1的基因敲低，其对照细胞系则表达无序shRNA对照序列。提取各组细胞的总RNA后，在Affymetrix微阵列上完成杂交实验。每个细胞系组别均设置3次生物学重复。