中文摘要：

microRNA-124（miR-124）丰富表达于脑组织中，调控神经细胞分化，同时在肿瘤组织中，miR-124还参与调控肿瘤细胞的增殖。分析表明，猪miR-124的前体序列位于4号和14号染色体中。本研究从长白猪（Sus scrofa）DNA基因组中扩增miR-124的前体序列，连接到T载体中，再经过EcoR Ⅰ和NotⅠ双酶切T载体，回收相应的酶切片段，将酶切片段连接到PiggyBac（PB）转座子载体中，然后将重组载体（PB-miR-124a1, PB-miR-124a2）转染猪肾细胞系PK15细胞，通过绿色荧光蛋白的表达检测转染效率，用嘌呤霉素进行稳转细胞的筛选，最后通过实时荧光定量PCR（qRT-PCR）测定细胞内miR-124的表达活性。结果显示，成功构建了重组转座子载体PB-miR-124a1和PB-miR-124a2，观察绿色荧光蛋白的表达，显示转染效率较高；稳定筛选的最佳嘌呤霉素的浓度为2 μg/mL。qRT-PCR结果显示，和对照组相比，稳定转染PB-miR-124a2组miR-124的表达显著升高（P＜0.01），而转染PB-miR-124a1组miR-124的表达升高不显著（P=0.06），说明miR-124的本底表达主要来源于ssc-mir-124a2基因。本实验为研究猪miR-124功能机制提供了基础材料。

英文摘要：

microRNA-124（miR-124）, high expression in brain, regulate neuronal differentiation and the proliferation of cancer cells. The porcine miR-124 precursor is transcribed from chromosome 4 and 14. In this study, the miR-124 precursor was amplified by PCR from landrace genomic DNA, ligated into T vector, then digested by EcoR Ⅰand NotⅠ, the digested fragment were connected into PiggyBac（PB） transposon vector, then the recombinant vector （PB-miR-124a1, PB-miR-124a2） was transfected to procine kidney epithelial cells（PK15）. The transfection efficiency was detected by expression of green fluorescent protein. The stable transfection cells were screened by puromycin, and the expression of miR-124 was determined by qRT-PCR. The results showed that the PB transposon recombinant vector （PB-miR-124a1, PB-miR-124a2） were constructed successfully with high transfection efficiency. The best stability screening concentration of puromycin was 2 μg/mL. qRT-PCR results showed that the expression of miR-124 in transfected PB-miR-124a2 group was increased significantly compared with control groups （P＜0.01）, but in transfected PB-miR-124a1 group, the expression of miR-124 increased not significantly （P=0.06）. These results demonstrated that the basal level of expression of miR-124 mainly transcribed from ssc-mir-124a2 gene. The study provides a foundation material for porcine miR-124 function research.