Leisy Douglas J, Lewis Teresa D, Leong Jo-Ann C, Rohrmann George F
Department of Microbiology, Oregon State University, 220 Nash Hall, Corvallis, OR 97331-3804, USA.
Hawaii Institute of Marine Biology, Kaneohe, HI 96744, USA.
J Gen Virol. 2003 May;84(Pt 5):1173-1178. doi: 10.1099/vir.0.18861-0.
Five fish cell lines were tested for their ability to be transduced by Ac-CAlacZ, a recombinant baculovirus that is capable of expressing a beta-galactosidase reporter gene from the CAG promoter (consisting of a cytomegalovirus enhancer element, a chicken actin promoter and rabbit beta-globin termination sequences). TO (Tilapia ovary), EPC (carp), CHH-1 (Chum salmon heart fibroblast) and CHSE-214 (chinook salmon embryo) cells were transducible, as demonstrated by an in situ beta-galactosidase assay, whereas RTG-2 (rainbow trout gonad) cells were not. The EPC cell line was used for more detailed studies on baculovirus transduction. The transduction frequency was found to be higher at 28 degrees C than at 21 degrees C. Addition of the histone deacetylase inhibitor sodium butyrate increased the number of blue cells detected 5- to 7-fold. The m.o.i. was positively correlated with transduction frequency, although the relationship did not appear to be strictly linear, as has been observed with mammalian cells. The temperature at which baculoviruses were adsorbed to EPC cells did not affect levels of beta-galactosidase expression. We also examined expression levels of beta-galactosidase in EPC cells after infection with a baculovirus construct that overexpresses the vesicular stomatitis virus G protein and displays it on the virion surface. Expression levels with this virus were approximately 15-fold higher than were observed with Ac-CAlacZ.
对五种鱼类细胞系进行了测试，以检测它们被Ac-CAlacZ转导的能力。Ac-CAlacZ是一种重组杆状病毒，能够从CAG启动子（由巨细胞病毒增强子元件、鸡肌动蛋白启动子和兔β-珠蛋白终止序列组成）表达β-半乳糖苷酶报告基因。通过原位β-半乳糖苷酶测定证明，罗非鱼卵巢（TO）细胞、鲤鱼上皮瘤（EPC）细胞、大麻哈鱼心脏成纤维细胞（CHH-1）和奇努克鲑胚胎（CHSE-214）细胞是可转导的，而虹鳟性腺（RTG-2）细胞则不可转导。EPC细胞系用于对杆状病毒转导进行更详细的研究。发现转导频率在28℃时高于21℃。添加组蛋白脱乙酰酶抑制剂丁酸钠可使检测到的蓝色细胞数量增加5至7倍。感染复数（m.o.i.）与转导频率呈正相关，尽管这种关系似乎不像在哺乳动物细胞中观察到的那样严格呈线性。杆状病毒吸附到EPC细胞上的温度不影响β-半乳糖苷酶的表达水平。我们还检测了用一种过表达水泡性口炎病毒G蛋白并将其展示在病毒粒子表面的杆状病毒构建体感染后，EPC细胞中β-半乳糖苷酶的表达水平。这种病毒的表达水平比用Ac-CAlacZ观察到的高约15倍。
