拟南芥AtXCD1蛋白的原核表达研究Prokaryotic Expression of Arabidopsis AtXCD1 Fusion Proteins
陈光朗;董万春;韩娇;樊婷婷;杨立波;曹树青;
摘要(Abstract):
[目的]研究拟南芥AtXCD1蛋白的原核表达。[方法]构建XCD1原核表达载体,根据NCBI基因序列设计引物,并以PCR扩增得到大量目的片段XCD1,将XCD1连接到原核表达载体pET-32a+,并对重组质粒进行测序鉴定,将重组质粒转化至大肠杆菌原核表达菌株BL21(DE3),对蛋白表达条件:诱导时间、诱导温度和IPTG浓度等进行优化。IPTG诱导表达获得目的蛋白,对蛋白表达条件:诱导时间、诱导温度和IPTG浓度等进行优化。[结果]XCD1序列全长为1 242 bp,与PCR产物大小一致。蛋白XCD1-pET-32a+较适表达条件为在30℃0.1 mmol/L的IPTG诱导1.5 h。[结论]该结果为进一步蛋白纯化及酶活测定试验奠定了基础。
关键词(KeyWords): 拟南芥(Arabidopsis thaliama);AtXCD1;重组载体;原核表达
基金项目(Foundation): 安徽省青年科学基金项目(1308085QC56);; 高等学校博士学科点基金(20120111110009)
作者(Author): 陈光朗;董万春;韩娇;樊婷婷;杨立波;曹树青;
Email:
DOI: 10.13989/j.cnki.0517-6611.2014.14.006
参考文献(References):
- [1]CLAUDIA JONAS,HIROFUMI NAKAGAMI,HERIBERT HIRT.Activation of distinct mitogen-activated protein kinase pathways by Copper and Cadmium[J].Plant Physiology,2004,136:3276-3283.
- [2]SHEORAN I S.Effect of cadmium and nicked on photosynthesis and the enzymes of the photosynthetic carbon reduction cycle in pigeonpea[J].Photosynthesis Research,1990,23:345-351.
- [3]侯文胜,郭三堆,路明,等.利用转基因技术进行植物遗传改良[J].生物技术通报,2002(1):10-15.
- [4]吴福彪.基因工程与植物的遗传改良[J].生物学通报,2010,45(5):7-10.
- [5]LOZANO-RODRIGUEZ E,HEMANDEZ L E,BONAY P,et al.Distribution of cadmiumin shoot and root tissues of maize and pea plants:physiological disturbances[J].Journal of Experimental Botany,1997,48:123-128.
- [6]NISHIZONO H,ICHIKAWA H,SUZIKI S,et al.The role of the root cell wall in the heavy metal tolerance of Athyrium yokoscense[J].Plant and Soil,1987,101:15-20.
- [7]吕申超,王海涛,鱼斌,等.拟南芥HIS1-3基因克隆及表达载体构建[J].合肥工业大学学报,2012(1):120-123.
- [8]CLAUDIA JONAK,HIROFUMI NAKAGAMI,HERIBERT HIRT.Heavy Metal Stress Activation of Distinct Mitogen-Activated Protein Kinase Pathways by Copper and Cadmium[J].Plant Physiology,2004,136:3276-3283.
- [9]SAMBROOK J,RUSSELL D W.分子克隆实验指南[M].黄培堂,王嘉玺,朱厚础,等,译.3版.北京:科学出版社,2002:26-395.
- [10]杜李继,顾菊,陈光朗,等.拟南芥AtMYB50和AtMYB61蛋白的原核表达研究[J].合肥工业大学学报,2014(1):105-109.