Isolation and Characterization of Matrix Attachment Regions from Green Alga:Dunaliella salina

Abstract

Matrix attachment region (MAR) is the DNA fragment that could bind to the nuclear matrix. Recently, many experiments have demonstrated that MAR can increase overall levels of foreign gene expression and decrease variability of expression. MARs have been isolated from yeast, plants, animals and human, but not from algae. In order to isolation the MAR from unicellular alga: Dunaliella salina, we first created a random MAR library as follows: The nuclear matrix were prepared from isolated nuclei by extracting with 25mM lithium diiodosalicylate (LIS) and then digested with restriction enzyme, the residual DNA fragments in the nuclear matrices were cloned into the pUC18 to construct the MAR library. Secondly, the interactions of several of the cloned DNA fragments with nuclear matrices were tested by an in vitro binding assay in which E.coli DNA was used as competitor. Some of DNA fragments could bind specifically to the nuclear matrices and four of them contained sequences related to known consensus motifs and a hairpin loop structure of MAR.