The Intron 1 Sequences Play the Role in Goat β-casein Gene Induced by Prolactin
| Autor: | Chun-Jen Chou, 周俊仁 |
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| Rok vydání: | 2011 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 99 Caseins, the major milk proteins, occupy 80% of total milk proteins. In mammary epithelial cells, the stage and tissue-specific expression of β-casein is under the induction of lactogenic hormone and extracellular matrix. There are several important transcription factor binding sites in β-casein proximal and distal promoter region, such as STAT5, C/EBP, GR, NF-1, Oct-1 and YY-1. These factors can affect the expression of β-casein. Many studies had indicated before, it could induce an increase of expression when the gene contained intron. We constructed the 5’-flanking 3.0 kb and 5.0 kb sequence of goat β-casein gene promoter into expression vectors. The 5.0 kb promoter vector contains complete intron 1 and exon 2 with signal peptide than 3.0 kb promoter vector. Results showed that the expression vector with exon 2 had the function of secretion and could be induced by lactogenic hormone. We observed the expression of the goat β-casein gene varied in NMuMG, and MCF-7 cell lines. The results indicated β-casein gene promoter activity possessed the specificity of cell species. According to references and sequence analyzed of intron 1 by using TFSEARCH and MOTIF Search to search for consensus sites in intron 1 sequence, we used electrophoretic mobility shift assay (EMSA) to analyze probe I (+16~+397) binding of intron 1 in presence of prolactin. The EMSA results showed that probe I (+16~+397) detect a clear band by prolactin induction. We predict +16~+397 of intron 1 might be important at gene transcription. And we used back to back PCR to delete probe I (+16~+397) region. From the results of +16~+397 delection vector, we confirmed that the expression decreased obviously in presence of prolactin induction. In coclusion, we found that probe I (+16~+397) region in goat β-casein gene intron 1 was able to induce by prolactin, and played an important role in β-casein gene. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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