Trastuzumab Resistance Breast Cancer Xenograft

Breast cancer cell line BT474 cells were administered subcutaneously into NCr nude mice. After randomization, mice received vehicle or trastuzumab (15mg/kg twice weekly). Tumors were categorized as control, responders, intrinsic resistance, and acquired resistance. RNA isolation was performed using RNeasy Plus kit (Qiagen) using manufacturer’s instructions. Sequencing libraries were created using Truseq stranded mRNA library preparation method from Illumina and sequenced on an Illumina NextSeq2000 sequencer. Reads were aligned to reference genome GRCh38 using HISAT2, then SAMTools was used sort and merge the alignments from multiple flow cells.将乳腺癌细胞系BT474的细胞皮下接种至NCr裸小鼠体内。经随机分组后，小鼠分别接受溶剂对照或曲妥珠单抗（trastuzumab，15mg/kg，每周2次）处理。将肿瘤样本划分为对照组、应答组、固有耐药组与获得性耐药组。总RNA提取采用Qiagen公司的RNeasy Plus试剂盒（RNeasy Plus kit），严格按照制造商的操作说明书执行。测序文库构建采用Illumina的Truseq链特异性mRNA文库制备方法，并在Illumina NextSeq2000测序仪上完成测序。将测序读段（reads）比对至参考基因组GRCh38时使用HISAT2工具，随后通过SAMTools对多个流动槽（flow cell）的比对结果进行排序与合并。