Transient reporter gene expression in oocysts and sporozoites of Cryptosporidium parvum controlled by endogenous promoters.
The apicomplexan protozoan Cryptosporidium parvum is an enteric parasite that affects a variety of mammal hosts including humans, and causes serious diarrheal disease in immunocompromised individuals, notably AIDS patients. Despite many advances in the development of transgenic techniques in many protozoan parasites over the past two decades, rare reports have been documented on the genetic manipulation on C. parvum. Achievement of the DNA-based transfection chiefly depends on the selection of an effective parasite genus-specific promoter. This report described the successful yellow (YFP-YFP) or red (RFP) fluorescent protein expression in oocysts and sporozoites of C. parvum controlled by the endogenous promoters of actin, alpha tubulin, and myosin genes using the restricted enzyme-mediated integration technique. One expression cassette in pBluescript backbone, YFP-YFP or RFP fused between 5' and 3' untranslated regions of actin gene, displayed the highest transfection efficiency with fluorescence rate around 50%. The established DNA-based transient transfection assay may contribute to a better understanding of the biology of Cryptosporidium species and their relationship with hosts and may also result in the development of more efficient molecule-based vaccines and drugs.
Key Laboratory of Zoonosis Research, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, China; College of Veterinary Medicine, Northeast Agricultural University, 59 Mucai Street, Harbin 150030, China.
Wei
W
Li
Key Laboratory of Zoonosis Research, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, China.
Yumei
Y
Diao
Key Laboratory of Zoonosis Research, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, China.
Pengtao
P
Gong
Parasitology Laboratory, College of Veterinary Medicine, China Agricultural University, Beijing 100193, China.
Xun
X
Suo
Key Laboratory of Zoonosis Research, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, China. Electronic address: jianhuali7207@163.com.
Jianhua
J
Li
Key Laboratory of Zoonosis Research, Ministry of Education, College of Veterinary Medicine, Jilin University, 5333 Xi'an Road, Changchun 130062, China. Electronic address: xczhang@jlu.edu.cn.
Xichen
X
Zhang
D019677:Artificial Gene Fusion; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D016785:Cryptosporidium parvum / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression; D017930:Genes, Reporter; D008164:Luminescent Proteins / Q000096:biosynthesis* / Q000235:genetics; D008967:Molecular Biology / Q000379:methods; D033661:Oocysts / Q000378:metabolism; D030781:Organisms, Genetically Modified; D010274:Parasitology / Q000379:methods; D011401:Promoter Regions, Genetic; D034101:Sporozoites / Q000378:metabolism; D000097573:Red Fluorescent Protein
D001426:Bacterial Proteins; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein; C054562:yellow fluorescent protein, Bacteria
Apicomplexa; Cryptosporidium parvum; Red fluorescent protein; Transfection; Yellow fluorescent protein