Aberrant products of mariner excision that have an impaired ability to be mobilized often include internal deletions that do not encroach on either of the inverted repeats. Analysis of 13 such deletions, as well as 7 additional internal deletions obtained by various methods, has revealed at least three internal regions whose integrity is necessary for efficient mariner mobilization. Within the 1286-bp element, the essential regions are contained in the intervals bounded by coordinates 229-586, 735-765, and 939-1066, numbering in base pairs from the extreme 5' end of the element. These regions may contain sequences that are necessary for transposase binding or that are needed to maintain proper spacing between binding sites. The isolation of excision-defective elements with point mutations at nucleotide positions 993 and 161/179 supports the hypothesis of sequence requirements, but the reduced mobility of transformation vectors with insertions into the SacI site at position 790 supports the hypothesis of spacing requirements. The finding of multiple internal regions that are essential for efficient mariner mobilization in vivo contrasts with reports that mini-elements with as little as 43 bp of DNA between the inverted repeats can transpose efficiently in vitro.