Using high-resolution annotation of insect mitochondrial DNA to decipher tandem repeats in the control region

Precise annotation of tick mitochondrial genomes reveals multiple STR variation and one transposon-like element

“…The 247 discovery of hybrid units suggested that mitochondrial DNA recombination can occur within an individual 248 tick, resulting in the insertion of [R28]m into [R34]l+n. This confirmed the identification of DNA 249 recombination events in our previous study of the E. fullo mitochondrial genome [5]. In that previous study,…”

“…The repeat 231 units in R1 were reverse complements to those in R2 (Figure 3B). Our DNA-seq data showed that the copy 232 numbers of R1 and R2 exhibited great diversity within an individual, which confirmed the finding from our 233 previous study of the E. fullo mitochondrial genome [5]. As repeat units in R1 and R2 were reverse 234 complements, we used PCR amplification (Table 1) combined with Sanger sequencing to further investigate 235 R1 sequences in more than 100 individual ticks from five species (D. silvarum, D. nuttalli, D. marginatus, D. 236 niveus, and D. ushakovae) and obtained the following results: (1) for each individual tick, the R1 sequence 237 obtained using Sanger sequencing is actually a consensus sequence of a large number of heterogeneous 238 sequences; (2) copy numbers were distributed between 2 and 5 for all studied repeat units, with one partial 239 repeat unit counted as 1; (3) in total, three types of repeat units of R1 with lengths of 28, 34, and 44 bp 240 (types 1, 2. and 3, respectively) were identified (Figure 3B) and noted as R28, R34 and R44; (4) in general, R1 241 sequences from ticks of one species were composed of repeat units of one type and R1 sequences from ticks 242 of the same species from different places had different copy numbers; and (5) among all studied ticks, D.…”

“…In this study, 111 we aimed to achieve the following research goals: (1) to provide a simple, cost-effective and accurate 112 method for the study of extremely high AT-content mitochondrial genomes within an individual animal 113 containing miniscule DNA. (2) to provide a high-quality reference genome with precise annotation for 114 future studies of tick mitochondrial genomes; and (3) to use these mitochondrial genomes containing two 115 Control Regions (CRs) to confirm our previous findings [5] from those containing one CR. 116 117 Results…”

“…In addition, we proposed that although all antisense transcripts are processed from two 93 primary transcripts, long antisense transcripts degrade quickly as transient RNAs, making them unlikely to 94 perform specific functions [4]. The second contribution involved the use 5′ and 3′ end small RNAs (5′ and 3′ 95 sRNAs) [4] to annotate mitochondrial genes at 1 bp resolution, subsequently dubbed precise annotation [5]. 96…”

“…Using these accurate genomes with precise annotations, we discovered that a novel 31-nt ncRNA exists in 97 mammalian mitochondrial DNA [4] and that the copy numbers of tandem repeats exhibit great diversity 98 within an E. fullo individual [5]. Recently, precise annotation of human, chimpanzee, rhesus macaque and 99 mouse mitochondrial genomes has been performed to investigate five Conserved Sequence Blocks (CSBs) 100 in the mitochondrial D-loop region [6], which ultimately led to an understanding of the mechanisms 101 involved in the RNA-DNA transition and even the functions of the D-loop.…”