Pooled-matrix protein interaction screens using Barcode Fusion Genetics.
Pooled-matrix protein interaction screens using Barcode Fusion Genetics.
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DOI：
10.15252/msb.20156660
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发表时间：
2016-04-22
影响因子：
9.9
通讯作者：
Roth FP
中科院分区：
文献类型：
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作者：
Yachie N;Petsalaki E;Mellor JC;Weile J;Jacob Y;Verby M;Ozturk SB;Li S;Cote AG;Mosca R;Knapp JJ;Ko M;Yu A;Gebbia M;Sahni N;Yi S;Tyagi T;Sheykhkarimli D;Roth JF;Wong C;Musa L;Snider J;Liu YC;Yu H;Braun P;Stagljar I;Hao T;Calderwood MA;Pelletier L;Aloy P;Hill DE;Vidal M;Roth FP
高通量二元蛋白质相互作用图谱正在继续扩展我们对细胞功能和疾病机制的理解。然而，我们距离人类和其他主要模式生物的完整相互作用图还有一到两个数量级的距离。完成这项工作将需要在更大的规模上使用许多互补的检测方法进行筛选，需要在蛋白质组规模的相互作用图谱中进一步提高效率。在这里，我们报告了Barcode Fusion Genetics‐Yeast Two‐Hybrid（BFG‐ Y2 H），通过该方法可以在单个多重菌株池中筛选完整的蛋白质对矩阵。BFG‐ Y2 H使用Cre重组融合来自不同质粒的DNA条形码，生成可通过下一代测序定量的嵌合蛋白质对条形码。我们将BFG-Y2 H应用于四种不同的基质，规模从~25 K到2.5 M蛋白质对。结果表明，BFG‐ Y2 H提高了蛋白质基质筛选的效率，质量与最先进的Y2 H方法相当。
High‐throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome‐scale interaction mapping. Here, we report Barcode Fusion Genetics‐Yeast Two‐Hybrid (BFG‐Y2H), by which a full matrix of protein pairs can be screened in a single multiplexed strain pool. BFG‐Y2H uses Cre recombination to fuse DNA barcodes from distinct plasmids, generating chimeric protein‐pair barcodes that can be quantified via next‐generation sequencing. We applied BFG‐Y2H to four different matrices ranging in scale from ~25 K to 2.5 M protein pairs. The results show that BFG‐Y2H increases the efficiency of protein matrix screening, with quality that is on par with state‐of‐the‐art Y2H methods.