SHORT PAPER
msechBari, a new MITE-like element in Drosophila sechellia
related to the Bari transposon

ELAINE SILVA DIAS AND CLAUDIA MARCIA APARECIDA CARARETO*
Department of Biology, UNESP – São Paulo State University, São José do Rio Preto, São Paulo, Brazil

(Received 18 August 2011; revised 2 November 2011; accepted 8 November 2011)

Summary

A few occurrences of miniature inverted-repeat transposable elements (MITEs) have been reported in species of
the genus Drosophila. Here, we describe msechBari, a MITE-like element in Drosophila sechellia. The element is
short, approximately 90 bp in length, AT-rich and occurs in association with, or close to, genes, characteristics
that are typical for MITEs. The identification was performed in silico using the sequenced genome ofD. sechellia
and confirmed in a laboratory strain. This short element is related to the Bari_DM transposon of Drosophila
melanogaster, having terminal inverted repeats (TIRs) of a similar length and a high identity with the full-length
Bari_DM element. The estimated recent origin of the element and the homogeneity observed between copies
found in the genome suggests that msechBari could be active in D. sechellia.

1. Introduction

Miniature inverted-repeat transposable elements
(MITEs) are non-autonomous short repeats that
mobilize within the host genome even without the
potential to encode the protein (i.e. the transposase)
responsible for their mobilization. The MITEs are, in
general, derived from ancient, related autonomous
elements, and their origin can occur through internal
deletions in autonomous elements, where the only
remaining are the terminal inverted repeats (TIRs)
and, sometimes, portions between the TIRs and the
coding region of the transposase. This origin supports
the proposal of their mobilization in trans by a trans-
posase encoded by a full-length element (Feschotte &
Pritham, 2007). The autonomous transposons use the
cellular machinery of the host cells for the protein
synthesis necessary for their mobilization, whereas the
MITEs use the machinery encoded by transposons
for mobilization. In the 1980s, Orgel & Crick (1980)
referred to ‘selfish DNA’ as the ‘ultimate parasites ’
due to the relationship of parasitism between auton-
omous elements and the machinery of the cell.
Recently, González & Petrov (2009) enlarged this idea

to include the MITEs because of their dependency on
autonomous elements for mobilization.

In general terms, MITE-like elements have been
widely described in plants (Moreno-Vazquez et al.,
2005; Lin et al., 2006; Guermonprez et al., 2008)
and specifically in grapevine (Benjak et al., 2009),
maize (Bureau & Wessler, 1992; Zerjal et al., 2009),
cereal grasses (Bureau & Wessler, 1994), Arabidopsis
(Feschotte & Mouches, 2000), rice (Feschotte et al.,
2003; Jiang et al., 2003; Nakazaki et al., 2003; Shan
et al., 2005), Medicago (Grzebelus et al., 2007, 2009),
apple (Han & Korban, 2007), beet (Menzel et al.,
2006), barley (Lyons et al., 2008; Petersen & Seberg,
2009), grasses (Park et al., 2003), pearl millet
(Remigereau et al., 2006) and pome fruit trees
(Wakasa et al., 2003). Descriptions in other organ-
isms, such as bacteria (Chen et al., 2008), cyano-
bacteria (Zhou et al., 2008), fungi (Xu et al., 2010),
silkworms (Han et al., 2010), fish (de Boer et al., 2007)
and amphibians (Hikosaka et al., 2011) are also found
in the literature, but few occurrences have been re-
ported in the Drosophila genus (Tudor et al., 1992;
Miller et al., 2000; Ortiz et al., 2010). Although nu-
merous MITEs have been identified, the association
with autonomous elements is often absent. Here, we
describe an MITE-like element found in the genome
of Drosophila sechellia that is associated with the Bari
transposon described inDrosophila melanogaster. The

* Corresponding author: Department of Biology, UNESP – São
Paulo State University, São José do Rio Preto, São Paulo, Brazil.
E-mail : carareto@ibilce.unesp.br

Genet. Res., Camb. (2011), 93, pp. 381–385. f Cambridge University Press 2011 381
doi:10.1017/S0016672311000371



high similarity found with Bari_DM in both TIRs and
internal regions suggests a close relationship with
autonomous elements.

2. Materials and methods

Searches for Bari_DM elements in the genomes of
species of Drosophila (unpublished data) resulted in
the identification of an y90 bp sequence, with TIRs
and no coding sequence, in the D. sechellia genome.
After this observation, the sequence of the TIRs of the
Bari_DM element of D. melanogaster (X67681) was
used to search the genome of D. sechellia (release 1.3,
June 2009) (Drosophila 12 Genomes Consortium,
2007) using the BLASTn software (Altschul et al.,
1990). Analyses aimed at identifying the target site
duplications (TSDs) and estimations of the gene
density in the adjacent regions of the MITEs were also
performed extracting the 10 kb 5k and 3k flanking re-
gions of each insertion. The ability to form secondary
structure was analysed using Mfold (Zuker, 2003)
(available at http://mfold.rna.albany.edu/).

To confirm that these MITEs were not a sequenc-
ing artefact, their occurrence was searched in a
D. sechellia strain maintained in our laboratory.
Genomic DNA was extracted from 50 individuals
according to a previously described protocol (Jowett,
1986). The amplification, cloning and sequencing
were performed using specific primers based on the
consensus sequence of the MITE identified in the
D. sechellia genome (Forward, 5k-MYRGTCAT-
GGTCAAAATTATTTTCACAA-3k and Reverse, 5k-
ACAGAGGTGGTCAAAAGTATTTTCACWW-3k).
PCR amplification was performed using 0.3125 unit
of Taq polymerase (Invitrogen), 200 ng genomic
DNA, 1 mM of MgCl2, 1rbuffer, 0.08 mM of dNTPs
and 0.4 mM of primers for a final volume of 25 ml. The
PCR conditions were as follows: initial denaturation
(94 xC, 120 s), followed by 30 cycles of denaturation
(94 xC, 15 s), annealing (59 xC, 10 s) and extension
(72 xC, 20 s). The PCR products were purified (DNA
GFX DNA & Gel Band, GE) and cloned (TOPO TA
Cloning kit, Invitrogen) according to the specifica-
tions of the manufacturers. Eight clones were selected
for extraction of the plasmid by a phenol/chloroform
protocol and sequenced using universal primers,
M13F and M13R, resulting in four sequences with
good quality.

The evolutionary relationships between the se-
quences were reconstructed using the software
Network with the Median Joining algorithm (Bandelt
et al., 1999) and the default parameters, using the
nucleotide sequences extracted from the D. sechellia
genome. The age of these insertions was estimated
using the following molecular clock equation (r=
k/2T), where r is the neutral synonymous substitution
rate of the Drosophila genus (r=0.011/site/Myr)

(Tamura et al., 2004) and k is the divergence rate
(Kimura 2-parameter distance) (Kimura, 1980). The
consensus sequence was reconstructed using the soft-
ware, DAMBE (Xia & Xie, 2001), and the distances
were calculated usingMEGA version 5 (Tamura et al.,
2011).

3. Results and discussion

In general, MITEs are smaller than 600 bp in length,
have conserved TIRs, a target site preference, no
coding potential and are AT-rich (Feschotte et al.,
2002). We found 49 MITE-like sequences in the se-
quenced genome of D. sechellia (see Supplementary
Table S1 available at http://journals.cambridge.org/
GRH) that presented lengths between 65 and 89 bp,
TIRs of 28 bp and AT contents of approximately
66%. Approximately 63% of these sequences are
flanked by AT dinucleotides, which are typical TSDs
of the MITE family Stowaway (Feschotte et al., 2002).
Both consensus sequences showed potential to form
secondary structure (see Supplementary Figure S1
available at http://journals.cambridge.org/GRH),
ability present in MITEs. Additionally, as other
MITEs (Zerjal et al., 2009; Han et al., 2010), these
sequences are preferentially associated with gene re-
gions (62% of the insertions were localized within
genes or harboured genes in their 10 kb flanking re-
gions).

The MITE-like sequences described here (Fig. 1
and Supplementary Table S1) show a high similarity
with the Bari_DM transposon described inD. melano-
gaster, but they are significantly smaller (65–89 bp)
than this autonomous element (1728 bp). Two types
of sequences were found, with their TIRs 100 and
89% similar to the Bari_DM, and both shared three
internal regions of 100% identity to Bari_DM and
between them. Thus, we concluded that the sequences
described in the D. sechellia genome are derivatives
of the Bari element, hereafter termed msechBari
elements.

These two types of msechBari, which essentially
differ by three nucleotides in their TIRs, were grouped
into two well-defined clusters in a network tree ; thus,
they can be considered to be two MITE subfamilies
(Fig. 2). The network suggests the existence of a
master sequence that would have given rise to the two
groups of sequences. Evolution under the master gene
model is characterized, in graphic reconstructions of
evolutionary relationships, by a star topology, where
the central sequence gives rise to the derived se-
quences (Cordaux et al., 2004). The length of the
branches is related to the elapsed time since the origin
of each sequence: short branches suggest a recent
origin, and long branches indicate an old origin.

The two subfamilies derived from the two master
sequences, msechBari1 and msechBari2, have

E. S. Dias and C. M. A. Carareto 382



short evolutionary distances within the group,
0.00341¡0.00036 and 0.00279¡0.0004, respectively ;
however, when a comparison was made between
the subfamilies, the distance was larger, 0.05020¡
0.00029. The short distances between the sequences
within a subfamily, the short branches and the ab-
sence of reticulation in the network suggest a recent
burst of transposition of these elements in the genome
of the strain that was sequenced. Accordingly, the
groupings of sequences in the network, represented by
large circles, indicate that the sequences are identical ;
therefore, these sequences are very recent and have
not had sufficient time to diverge. Similar events have
been reported for other transposable elements (Yang
et al., 2006; de Boer et al., 2007; Marzo et al., 2008;

Konovalov et al., 2010; Lerat, 2010) and MITE-like
sequences in different organisms (Jiang et al., 2003;
Chen et al., 2008; Zhou et al., 2008; Han et al., 2010;
Hikosaka et al., 2011). This recent origin is also sup-
ported by the average time of origin of the insertions
of each subfamily, 155 000 years (msechBari1) and
127 000 years (msechBari2).We confirmed the presence
of msechBari in a laboratory strain (Fig. 1). The se-
quences found were similar to those in the D. sechellia
sequenced genome. They had the internal region
conserved, but the 2 bp of the 5k TIRs were variable
(see Supplementary Figure S2 available at http://
journals.cambridge.org/GRH). This variation, if real,
could indicate inactivity of these MITES. However,
as we obtained only four sequences, it is possible that

Fig. 1. Alignment of the MITE-like sequences identified in the sequenced genome of D. sechellia with the consensus
sequence of the transposon, Bari_DM, as described in D. melanogaster. The shaded region corresponds to the TIRs, with
the three diagnostic substitutions of the two MITE subfamilies highlighted with asterisks; the boxes indicate the remaining
non-coding regions found in the MITEs and the dotted region corresponds to the not shown nucleotides 77–1444 present
only in the transposon Bari_DM.

Fig. 2. Network of the MITE-like sequences identified in the sequenced genome of D. sechellia. The size of the circles
corresponds to sequence frequency; the size of the branches is proportional to the number of mutations occurred, as
indicated by numbers above branches. Black circles correspond to the sequences of msechBari1 subfamily and the grey
circles to the msechBari2 subfamily.

MITEs in D. sechellia 383



the two pairs of variable bases are sequencing arte-
facts.

For the mobilization of a transposon, such as Bari,
the transposase proteins recognize and bind to specific
sites in the TIRs to promote transposition. For some
MITEs found in plants, the mobilization of trans-
posons that do not have coding capacity has been
suggested to occur via transposases in trans from
elements that are distantly related. For example, up to
approximately 20 000 insertions of rice MITE-like
elements of the Stowaway family, which exhibit TIRs
similar to other mariner-like elements, have been re-
ported. However, these elements are not homologous
to any other autonomous elements that have been
described in rice ; thus, it has been proposed that these
elements be mobilized by a transposase encoded
by other distantly related autonomous elements
(Feschotte, 2008). Therefore, the recent transposition
of the msechBari could have resulted from the pres-
ence of a transposase from an active Bari_DM trans-
poson in D. sechellia or from other Bari-like elements
in the D. sechellia genome that can recognize the
TIRs. Only two full-length Bari copies, with both in-
tact TIRs, were found in the D. sechellia genome, but
both have many stop codons in their transposase
coding sequences (see Supplementary Figure S3
available at http://journals.cambridge.org/GRH), in-
dicating that the Bari element in D. sechellia is in-
active. However, both copies exhibit a low diversity,
when compared with the consensus sequence of
Bari_DM, suggesting that this inactivity is recent.
Therefore, this autonomous element is potentially re-
sponsible for the msechBari mobilization in the recent
past ; however, the mobilization by another distantly
related autonomous element cannot be disregarded.

Funding for this project was provided by the Brazilian
agencies, FAPESP – Fundação de Amparo à Pesquisa do
Estado de São Paulo (Grant 2010/10731-4 to C. M. A. C.
and fellowship 2008/07629-3 to E. S. D.), CNPq (Conselho
Nacional de Desenvolvimento Cientı́fico e Tecnológico)
(Grant 304880/2009-4 to C. M. A. C.) and FUNDUNESP
(Fundação para o Desenvolvimento da UNESP) (Grant
670/10). We thank Jean David, PhD for providing the strain
used in this study.

4. Supplementary material

The online data are available at http://journal-
s.cambridge.org/GRH

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