END JOINING OF GENOMIC DNA AND TRANSGENE DNA IN FERTILIZED MOUSE EGGS

A linear 5.2-kb HS2/beta-globin construct with an upstream KpnI termin us (4-nucleotide (nt) 3' protruding single strand, PSS) and a downstre am SalI terminus (4-nt 5' PSS) was microinjected into fertilized mouse eggs. The injected DNA fragments integrated into the mouse genome pri marily as a head-to-tail tandem array. Chromosome/transgene junctions were obtained from seven of eight transgenic animals. All of the junct ions occurred in the proximity of a transgene KpnI end; a maximum loss of 8 nt from the transgene terminus was observed. Two of these juncti ons completely preserved the 4-nt KpnI 3' PSS. Transgene/transgene jun ctions from two animals were analyzed. SalI/KpnI junctions that comple tely preserved both the SalI 5' PSS and the KpnI 3' PSS were found in each animal. These are the first examples of complete nt preservation at junctions formed between a 5' PSS terminus and a 3' PSS terminus in transgenic mice. The data are consistent with the fill-in model of Th ode et al. [Cell 60 (1990) 921-928] in which alignment proteins juxtap ose 5' PSS and 3' PSS termini; DNA polymerase then utilizes the recess ed 3'-OH of the 5' PSS terminus as a primer to synthesize DNA across t he gap. This mechanism results in the formation of junctions with no l oss of sequence. The results described in the present paper suggest th at this mechanism may be involved in the formation of junctions in tra nsgenic mice.