Development of an efficient targeted cell-SELEX procedure for DNA aptamer reagents.
Development of an efficient targeted cell-SELEX procedure for DNA aptamer reagents.
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开发有效的DNA适体试剂的有效靶向细胞 - 塞氏菌手术。
DOI：
10.1371/journal.pone.0071798
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Thomson JA
中科院分区：
文献类型：
--
作者：
Meyer S;Maufort JP;Nie J;Stewart R;McIntosh BE;Conti LR;Ahmad KM;Soh HT;Thomson JA
通过细胞-SELEX产生的DNA适体提供了抗体的有吸引力的替代方案，但是产生针对特定的已知膜蛋白靶标的适体已被证明具有挑战性，并且严重限制了适体作为用于细胞鉴定和纯化的亲和试剂的使用。我们修改了BJAB淋巴母细胞瘤细胞系过表达鼠c-kit细胞表面受体。经过六轮细胞-SELEX、高通量测序和生物信息学分析后，我们鉴定了结合表达c-kit的BJAB细胞但不结合野生型BJAB对照的适体。这些适体之一还识别由肥大细胞系或造血祖细胞内源性表达的c-kit，并特异性阻断c-kit配体干细胞因子（SCF）的结合。该适体能够通过荧光激活细胞分选（FACS）从混合的骨髓群体中更好地分离c-kit+造血祖细胞，这表明该方法可广泛用于快速分离适合于纯化其他特定细胞类型的亲和试剂。在这里，我们描述了一种新的程序，有效地产生DNA适体，结合到特定的细胞膜蛋白，并可用作高亲和力试剂。我们将该过程命名为STACS（Specific TTarget Cell-SELEX）。
DNA aptamers generated by cell-SELEX offer an attractive alternative to antibodies, but generating aptamers to specific, known membrane protein targets has proven challenging, and has severely limited the use of aptamers as affinity reagents for cell identification and purification. We modified the BJAB lymphoblastoma cell line to over-express the murine c-kit cell surface receptor. After six rounds of cell-SELEX, high-throughput sequencing and bioinformatics analysis, we identified aptamers that bound BJAB cells expressing c-kit but not wild-type BJAB controls. One of these aptamers also recognizes c-kit endogenously expressed by a mast cell line or hematopoietic progenitor cells, and specifically blocks binding of the c-kit ligand stem cell factor (SCF). This aptamer enables better separation by fluorescence-activated cell sorting (FACS) of c-kit+ hematopoietic progenitor cells from mixed bone marrow populations than a commercially available antibody, suggesting that this approach may be broadly useful for rapid isolation of affinity reagents suitable for purification of other specific cell types. Here we describe a novel procedure for the efficient generation of DNA aptamers that bind to specific cell membrane proteins and can be used as high affinity reagents. We have named the procedure STACS (Specific TArget Cell-SELEX).
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影响因子：
14.9
作者：
Goujon M;McWilliam H;Li W;Valentin F;Squizzato S;Paern J;Lopez R
通讯作者：
Lopez R
DOI：
10.3390/molecules15010001
发表时间：
2009-12-24
期刊：
Molecules (Basel, Switzerland)
影响因子：
--
作者：
Avci-Adali M;Paul A;Wilhelm N;Ziemer G;Wendel HP
通讯作者：
Wendel HP
选择针对未糖基化的EGFRVIII胞外域的适体，细胞内递送降低膜结合的EGFRVIII并诱导细胞凋亡。
影响因子：
3.7
作者：
Liu Y;Kuan CT;Mi J;Zhang X;Clary BM;Bigner DD;Sullenger BA
通讯作者：
Sullenger BA
影响因子：
64.5
作者：
Ding, S;Wu, XH;Xu, T
通讯作者：
Xu, T
影响因子：
14.9
作者：
Thiel KW;Hernandez LI;Dassie JP;Thiel WH;Liu X;Stockdale KR;Rothman AM;Hernandez FJ;McNamara JO 2nd;Giangrande PH
通讯作者：
Giangrande PH