Using labeled transcripts generated in vitro from squid total genomic DNA as a probe, we isolated and characterized a SINE that is present in the squid genome. The squid SINE appears to be derived from a tRNA(Lys). When the consensus sequences of five different SINEs with a tRNA(Lys)-like structure from distantly related species, including squid, were aligned, we found in the tRNA-unrelated region two sequence motifs that were almost identical among these five SINEs. This observation suggests a common evolutionary origin for these SINEs and/or some function(s) for these motifs. Similar sequences were unexpectedly found to be present in sequences complementary to the U5 regions of several mammalian retroviruses whose primer is a tRNA(Lys). On the basis of these findings, we present a model for the generation of SINEs. We propose that they are derived from a "strong-stop DNA" with a primer tRNA(Lys) that is an intermediate in the reverse transcription of certain retroviruses. Our model suggests that a certain group of SINEs may have been generated by horizontal transmission, although it is not clear whether information was transmitted via a similar retrovirus or via an RNA or DNA of a SINE.