Global skin gene expression profile of early diffuse cutaneous systemic sclerosis shows a prominent innate and adaptive immunity profile. Global skin gene expression profile of early diffuse cutaneous systemic sclerosis shows a prominent innate and adaptive immunity profile

RNA from skin biopsies from 48 patients in the Prospective Registry for Early Systemic Sclerosis (PRESS) cohort (mean disease duration 1.3 years) and 33 matched healthy controls was examined by nextGen RNA sequencing Overall design: 3- or 4-mm diameter punch biopsies were obtained from the forearm skin and immersed in RNAlater solution (Qiagen). RNA was extracted using miRNeasy Mini kits (Qiagen). cDNA libraries were prepared using the Illumina TruSeq stranded Total RNA Library Prep Gold kit, loaded on cBot (Illumina) at a final concentration of 10 pM to perform cluster generation, followed by 2 x 76 bp paired-end sequencing on HiSeq 2500 (Illumina), generating on average around 50 million reads per sample.

本研究对早期系统性硬化症前瞻性登记队列（Prospective Registry for Early Systemic Sclerosis, PRESS）中的48例患者（平均病程1.3年）及33例匹配健康对照的皮肤活检组织RNA，采用下一代RNA测序（nextGen RNA sequencing）技术进行了检测。实验整体设计如下：从前臂皮肤获取直径3或4毫米的穿刺活检样本，将其浸没于RNAlater保存液（Qiagen）中。使用miRNeasy Mini试剂盒（Qiagen）提取总RNA。采用Illumina TruSeq链特异性总RNA文库制备金试剂盒（Illumina TruSeq stranded Total RNA Library Prep Gold kit）构建cDNA文库，以10 pM的终浓度上样至cBot（Illumina）进行簇生成，随后在HiSeq 2500（Illumina）测序平台上开展2×76 bp双端测序，每个样本平均产生约5000万条reads。