The construction and characterization of the bi-directional promoter between pp38 gene and 1.8-kb mRNA transcripts of Marek's disease viruses.
BACKGROUND
Marek's disease virus (MDV) has a bi-directional promoter between pp38 gene and 1.8-kb mRNA transcripts. By sequencing for the promoters from 8 different strains (CVI988, 814, GA, JM, Md5, G2, RB1B and 648A), it is found, comparing with the other 7 MDV strains, CVI988 has a 5-bp (from -628 to -632) deletion in this region, which caused a Sp1 site destroyed. In order to analysis the activity of the promoter, the complete bi-directional promoters from GA and CVI988 were, respectively, cloned into pCAT-Basic vector in both directions for the recombinants pPGA(pp38)-CAT, pPGA(1.8 kb)-CAT, pPCVI(pp38)-CAT and pPCVI(1.8 kb)-CAT. The complete promoter of GA was divided into two single-direction promoters from the replication of MDV genomic DNA, and cloned into pCAT-Basic for pdPGA(pp38)-CAT and pdPGA(1.8 kb)-CAT as well. The above 6 recombinants were then transfected into chicken embryo fibroblasts (CEFs) infected with MDV, and the activity of chloramphenicol acetyltransferase (CAT) was measured from the lysed CEFs 48 h post transfection.


RESULTS
The results showed the activity of the divided promoters was decreased on both directions. In 1.8-kb mRNA direction, it is nearly down to 2.4% (19/781) of the whole promoter, while it keeps 65% (34/52) activity in pp38 direction. The deletion of Sp1 site in CVI988 causes the 20% activity decreased, and has little influence in pp38 direction.


CONCLUSIONS
The present study confirmed their result, and the promoter for the 1.8-kb mRNA transcripts is a much stronger promoter than that in the orientation for pp38.
Chen
Ruiai
R
College of Biological Sciences, China Agricultural University, Beijing 100193, China. chensa727@yahoo.com.cn
Ding
Jiabo
J
Wang
Bin
B
D000818:Animals; D000956:Antigens, Viral / Q000235:genetics*; D002478:Cells, Cultured; D002645:Chickens; D017353:Gene Deletion; D015967:Gene Expression Regulation, Viral; D025922:Mardivirus / Q000235:genetics*; D010750:Phosphoproteins / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics*; D014158:Transcription, Genetic
D000956:Antigens, Viral; D010750:Phosphoproteins; D012333:RNA, Messenger; C071698:phosphoprotein pp38, Marek's disease virus