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The evolutionary process by which functional gene copies become non-functional pseudogenes is of special importance because non-functional copies can be the source of functional genes in the genome. The most comprehensive study of this phenomenon in humans revealed that non-functional copies for both protein-coding and non-protein-coding genes are frequent in the genome and are mostly nuclear, suggesting that germ-line retrotransposition is the major cause of their origin [1]. Despite a wealth of studies on non-functional genes in the human lineage, little is known about the origin of non-functional retrogenes in non-human primates. Here, we collected full-length non-functional retrogenes from three representative non-human primates (human, old world monkey, and prosimian) as well as from two other old world monkeys (marmosets and squirrel monkeys), and we screened their flanking sequences to determine if they were retroposon-derived pseudogenes. We confirmed the presence of non-retroposon sequences in 36% of the retrogenes, which exhibited structural features that are characteristic of retrocopies including long stretches of repeats, partial pseudogenization, and chimeric genic features. Thus, the non-retroposon sequences of the 36% of the retrogenes appeared to have been generated by retroposon insertions. However, about half of them contained sequences related to the tRNA-derived sequences, suggesting that they were retroposon-derived pseudogenes. None of the reengineered sequences contained DNA fragments derived from the integrase coding gene or other structural hallmarks of retrocopies. Rather, they had chimeric genic features suggesting that they were the products of cDNA retrotransposition. Furthermore, while the origin of the non-retroposon sequences of 29 of these non-retroposon-derived retrogenes could be determined unambiguously, they were derived from tRNA

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