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Neutrophil gelatinase-associated lipocalin (neutrophil gelatinase associatedlipocalin, NAGL) is the lipocalin (lipocallin) family, a new member of the 1993 in human neutrophils was first discovered . Homology analysis of NGAL mouse oncogene product the 24 p3 homology ; another study found significantly increased in tumor tissue such as colon cancer and breast cancer , esophageal cancer cell lines and ovarian cancer cell lines in NGAL expression . This suggests that NGAL may be a new human oncogene . However, so far , NGAL overexpression in tumor cells regulation mechanism is unclear . To this end , this ' research combined use of PCR, DNA recombinant nested deletions , as well as dual - luciferase reporter gene detection technology positioning NGAL promoter region , while taking advantage of bioinformatics methods to predict the trans-acting factors for the elucidation of NGAL expression regulation mechanism to provide the basic data . Content and methods : 1 . Extracted SHEEC total cellular DNA as a template , using six pairs of primers for PCR reaction , get NGAL5 ' flanking region of different length fragments , -1431 - 84 -1 137 - 84 -945 - 84 -657 - 84 , -416 - 84 -152 - 84 . First, the fragment insert pGEM-Teasy, and then subcloned into pGl3-Basic, build luciferase reporter gene assay series plasmid pB1431 pB1137 pB945 , pB657 , pB416 and pB152 the approximate distribution segment identified NGAL promoter . Joint use of nested deletions and dual luciferase reporter gene assay technology , NGAL promoter further determine a more exact location . 3 . Use of bioinformatics methods to predict the NGAL promoter region functional components and its trans-acting factors to explore the mechanism of regulation of gene expression . Results : 1 . Compared with the empty vector pGL3-Basic pB1431 pB1137 , pB945 , pB657 , pB416 and pB152 such as relative luciferase activity were significantly increased (P lt; 0.01 or lt ; 0.05 ) Description NGAL start
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