Defining the Sequence Recognized with BmFTZ-F1, a Sequence Specific DNA Binding Factor in the Silkworm, Bombyx Mori, As Revealed by Direct Sequencing of Bound Oligonucleotides and Gel Mobility Shift Competition Analysis

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 1991 Jul 11

PMID 1852610

Citations 14

Authors

H Ueda

S Hirose

Affiliations

Soon will be listed here.

Abstract

BmFTZ-F1 is a Bombyx mori homologue of FTZ-F1, a positive regulator of the fushi tarazu gene of Drosophila melanogaster. In order to determine the sequence recognized with this factor, we made three sets of oligonucleotide mixture which contain 4 possible nucleotides at different positions within the previously proposed 12-bp binding consensus sequence. Oligonucleotides which bound to purified BmFTZ-F1 were separated by a gel mobility shift procedure and a binding sequence was determined by direct sequencing through Maxam-Gilbert method. By this analysis, 7 positions showed clear sequence preference and 5 positions showed weak or no sequence preference. The importance of each nucleotide at each position was confirmed by a gel mobility shift competition analysis and results were presented as a quantitative difference in the binding affinity. From these analyses, we conclude that the best binding sequence of BmFTZ-F1 is 5'-PyCAAGGPyCPu-3'. This method may be useful for the determination of a binding sequence of other sequence specific DNA binding factor.

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