小鼠SLC基因真核表达载体的构建及其趋化活性鉴定

目的：克隆小鼠次级淋巴样组织趋化因子(secondary lymphoid-tissue chemokine，SLC)基因，并构建真核表达载体。在体内外检测该表达载体表达产物的免疫趋化功能。方法：用RT—PCR从C57BL／6小鼠胸腺组织中克隆小鼠SLC基因，构建真核表达载体pcDNA3．1—msLC。在体外，用基因枪转染小鼠黑色素瘤B16F10细胞；RT—PCR检测转染细胞有SLC的表达；利用趋化小室法，检测表达产物针对淋巴细胞的趋化活性。在体内，用基因枪通过小鼠皮肤局部转染SLC基因，观察转染局部淋巴细胞的浸润。结果：克隆的基因经测序证实，为小鼠SLC基因Scya21b型。转染SLC基因的B16F10细胞体外培养48h后，RT—PCR检测到SLC的表达，同时培养基上清具有针对淋巴细胞的趋化活性。通过基因枪局部转染小鼠皮肤，24h后皮肤病理检查显示，局部皮内有明显的淋巴细胞浸润。结论：从小鼠胸腺组织中克隆到小鼠SLC基因，构建的真核表达载体pcDNA3．1—mSLC在体内外均可以表达，并具有针对淋巴细胞的趋化活性。

Construction of eukaryotic expression vector of murine SLC gene and charac-terization of its chemotactic function

AIM: To construct the eukaryotic expression vector of mu-rine SLC gene and study the chemotactic function of murine SLC in vitro and in vivo. METHODS; Murine SLC gene was cloned by RT-PCR from the thymus tissue of a C57BL/6 mouse. Eukaryotic expression vector of SLC gene - pcD-NA3.1-mSLC was constructed and transfected into B16F10 cells by gene gun. Culture supernatant was collected 48 hours after the transfection and chemotactic function of expression product to lymphocytes was detected by a chemo-taxis chamber. SLC expression was detected by RT-PCR. Lymphocytic infiltration was observed in SLC gene transfected murine abdominal skin. RESULTS: The gene cloned from the C57BL/6 mouse thymus tissue was SLC gene Scya21b. Transfected cells expressed SLC mRNA, and culture supernatant of those cells had a potent chemotactic function to lymphocytes. Histological examination of transfected skin showed obvious lymphocytic infiltration. CONCLUSION : The SLC gene was cloned from the mouse thymus tissue and could be expressed in B16F10 cells. The expression product in vitro and in vivo, had a remarkable chemotactic function to lymphocytes.