Comparative analysis of complete mitochondrial DNA control region of four species of Strigiformes.

Abstract

The sequence of the whole mitochondrial (mt) DNA control region (CR) of four species of Strigiformes was obtained. Length of the CR was 3,290 bp, 2,848 bp, 2,444 bp, and 1,771 bp for Asio flammeus, Asio otus, Athene noctua, and Strix aluco, respectively. Interestingly, the length of the control region was maximum in Asio flammeus among all the avian mtDNA control regions sequenced thus far. In addition, the base composition and organization of mtDNA CR of Asio flammeus were identical to those reported for other birds. On the basis of the differential frequencies of base substitutions, the CR may be divided two variable domains, I and III, and a central conserved domain, II. The 3' end of the CR contained many tandem repeats of varying lengths and repeat numbers. In Asio flammeus, the repeated sequences consisted of a 126 bp sequence that was repeated seven times and a 78 bp sequence that was repeated 14 times. In Asio otus, there were also two repeated sequences, namely a 127 bp sequence that was repeated eight times and a 78 bp sequence that was repeated six times. The control region of Athene noctua contained three sets of repeats: a 89 bp sequence that was repeated three times, a 77 bp sequence that was repeated four times, and a 71 bp sequence that was repeated six times. Strix aluco, however, had only one repeated sequence, a 78 bp sequence that was repeated five times. The results of this study seem to indicate that these tandem repeats may have resulted from slipped-strand mispairing during mtDNA replication. Moreover, there are many conserved motifs within the repeated units. These sequences could form stable stem-loop secondary structures, which suggests that these repeated sequences play an important role in regulating transcription and replication of the mitochondrial genome.

Full text

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遗遗 传传 学学 报报 Acta Genetica Sinica, November 2006, 33 (11)：965–974 ISSN 0379-4172
Comparative Analysis of Complete Mitochondrial DNA Con-
trol Region of Four Species of Strigiformes
XIAO Bing1, MA Fei1, SUN Yi2, LI Qing-Wei1,①
1. College of Life Sciences, Liaoning Normal University, Dalian 116029, China;
2. School of Life Sciences, Xiamen University, Xiamen 361005, China
Abstract: The sequence of the whole mitochondrial (mt) DNA control region (CR) of four species of Strigiformes was obtained.
Length of the CR was 3 290 bp, 2 848 bp, 2 444 bp, and 1 771 bp for Asio flammeus, Asio otus, Athene noctua, and Strix aluco,
respectively. Interestingly, the length of the control region was maximum in Asio flammeus among all the avian mtDNA control
regions sequenced thus far. In addition, the base composition and organization of mtDNA CR of Asio flammeus were identical to
those reported for other birds. On the basis of the differential frequencies of base substitutions, the CR may be divided two variable
domains, Ⅰ and Ⅲ, and a central conserved domain, Ⅱ. The 3′ end of the CR contained many tandem repeats of varying lengths
and repeat numbers. In Asio flammeus, the repeated sequences consisted of a 126 bp sequence that was repeated seven times and a
78 bp sequence that was repeated 14 times. In Asio otus, there were also two repeated sequences, namely a 127 bp sequence that
was repeated eight times and a 78 bp sequence that was repeated six times. The control region of Athene noctua contained three sets
of repeats: a 89 bp sequence that was repeated three times, a 77 bp sequence that was repeated four times, and a 71 bp sequence that
was repeated six times. Strix aluco, however, had only one repeated sequence, a 78 bp sequence that was repeated five times. The
results of this study seem to indicate that these tandem repeats may have resulted from slipped-strand mispairing during mtDNA
replication. Moreover, there are many conserved motifs within the repeated units. These sequences could form stable stem-loop
secondary structures, which suggests that these repeated sequences play an important role in regulating transcription and replication
of the mitochondrial genome.
Key words: Strigiformes; mitochondrial DNA; control region (CR); tandem duplication; molecular evolution

Received: 2006-06-15; Accepted: 2006-07-17
This work was supported by the National Natural Sciences Foundation of China (No. 30470936).
① Corresponding author. E-mail: Liqw@263.com; Tel & Fax: +86-411-8582 7799
The control region (D-loop) is the only major
noncoding segment in the vertebrate mitochondrial
genome. It is also the most variable part of the
mtDNA and evolves three to five times more rapidly
compared with the rest of the mitochondrial ge-
nome[1]. In Aves, the mitochondrial control region is
located between the tRNAGlu gene and the tRNAPhe
gene[2], and its primary function is usually believed to
be the regulation of replication and transcription of
the mitochondrial genome[3]. In particular, sequence
variation of the control region may result in length
differences in bird mitochondrial genomes. In addi-
tion, many studies have shown that the extensive size
variation in the mtDNA control region in birds is at-
tributed to the insertion-deletion of some segments
and/or the variation of the copy number-length of
tandem repeat sequences within its 5′ and 3′ ends.
Therefore, a better understanding of the structural
property of the control region is important for studies
on avian phylogeny and population.
Over the past ten years, with the sequencing of
the mitochondrial control region of many Classes,
such as Cyclostomata, Pisces, Amphibia, Reptile,
Aves, and Mammalia, a great deal of attention has
been paid to the evolutionary mechanism of the con-
trol region and its application in molecular phyloge-
netic analysis. Currently, the mitochondrial control
region sequences of 22 avian species have been de-

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posited in the GenBank. However, there are only five
incomplete sequences from Strigiforme species in the
GenBank, namely, Strix occidentalis (DQ087169),
Strix aluco (DQ092611), Ninox novaeseelandiae
(AY309457), Strix nigrolineata (AY830859), and
Strix uralensis (AY836774). To further elucidate the
structural character of the complete mitochondrial
control region of Strigiformes, in this study, the se-
quence of the complete mtDNA control region of four
owls is reported and the base composition and the
organization of the control region is analyzed. Fur-
thermore, the origin and the evolution of tandem re-
peats of the mtDNA control region as well as their
function and application are discussed.
1 Materials and Methods
1. 1 Samples
Asio flammeus and Asio otus were collected
from the National Natural Protective Zone of Lao Tie
Hill, Dalian. Strix aluco and Athene noctua were col-
lected from Kunming, Yunnan Province. Fresh liver
tissue samples were extracted and frozen at −80.
℃
1. 2 DNA extraction
Mitochondrial DNA of the four owls was ex-
tracted from frozen liver tissue according to the pro-
cedure described by Arnason et al [5].
1. 3 DNA amplification and sequencing
Primers (Table 1) were designed for the four
owls. PCR products were sequenced using the meth-
ods of Long-PCR and primer walking. Complete
mtDNA control region including all adjacent seg-
ments were sequenced bidirectionally until overlap-
ping adjacent segments of approximately 80-120 bp
were obtained to ensure accuracy. All sequencing
reactions were carried out on an ABI 377 DNA
automated sequencer (PE Applied Biosystems).
1. 4 Sequence analysis
The locations of genes were determined by
comparisons with sequences of the mtDNA control
region of Gallus gallus (X52392), Ninox novaesee-
landiae (AY309457), and Buteo buteo (AF380305).
Clustal X 1.8[6] software, Sequencher software,
and DNASTAR software were used to analyze nu-
cleotide composition and sequence alignment to de-
termine the length of the control region, as well as to
search conserved sequence motifs. Potential secon-
dary structures of the nucleotide sequence of the con-
trol region were analyzed using RNAstructure4.2
program. MEME-Motif discovery tool was used to
analyze conserved motif in the repeated sequences.
2 Results
2. 1 The length of the control region
Sequencing results showed that the control re-
gion of Asio flammeus, Asio otus, Athene noctua, and
Strix aluco was 3 290 bp, 2 848 bp, 2 444 bp, and
1 771 bp long, respectively. Thus, the control region
of the four owls is much longer than that of other
birds (about 1-1.5 kb). It is also interesting that the
length of the control region is maximum in Asio
flammeus among all the avian mtDNA control regions
sequenced thus far.
2. 2 The organization of the control region
The base composition and organization of the

Table 1 Primers for amplification
Primers
1 S-Cytb-13354F
2 S-12S-256R
3 S-ND6-527F
4 S-Ctyb-221R
5 L-Ctyb-1592F
6 L-12S-211R
Note: 1 and 2 were the primers for Strix aluco and Athene noctua; 3 and 4 were the primers for Asio flammeus; 5 and 6 were the
primers for Asio otus.
Nucleotide sequence (5′→3′)
GCTGACTACTCCGCAACCTACACGCAAATG
GAATGTAGCCCATCTCTTCCACCTCATAGG
AAGCCGCCGTTAACTCACCC
GTGGTGGGAGCATTAGGATT
GGTGTAGGAGGGGGGAAA
CCTTGGGTGTTCTGTGGTGA
Derived source
This study
This study
This study
This study
This study
This study

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XIAO Bing et al.: Comparative Analysis of Complete Mitochondrial DNA Control Region of Four Species of Strigiformes 967


control region of the four owls are very similar to
those reported for other birds, i.e., low G content and
considerably AT-rich content. The nucleotide compo-
sition of the mtDNA control region of the four owls is
listed in Table 2.
On the basis of the differential frequencies of the
base substitutions, the mitochondrial control region
may be divided into three domains[7-9]: two variable
domains, Ⅰ and , and a central conserved domain,
Ⅲ
Ⅱ. The general structure of the mitochondrial DNA
control region of the four owls is shown in Fig. 1.
Some conserved sequence motifs are also found in all
the three domains, including ETAS-1, ETAS-2, and
CSB-1-like motif in domain
E-box, F-box, and bird similarity box in domain
and CSB-1 motif in domain Ⅲ[10]. However, the four
owls lack heavy strand replication origin OH and the
bidirectional transcription promoter LSP/HSP. Fur-
thermore, CSB-2 and CSB-3 motifs that have been
identified in some vertebrates have not been found in
the four owls. A comparison of the conserved se-
quence block is shown in Table 3. It is very clear that
the homologies of the conserved sequence blocks in
domain are highe
Ⅱ
r than those in domainsⅠ and
; B
Ⅰ
-box, C-box, D-box,
;
Ⅱ
,
Ⅲ
suggesting that domain
pared with domainsⅠ and
is more conservative co
.
Ⅲ
Ⅱ
m-
2. 3 Tandem duplication sequences
The 3′ end of the control region contains many
tandem duplication sequences. In Asio flammeus, the
repeated sequences consisted of a 126 bp sequence
that was repeated seven times and a 78 bp sequence
that was repeated 14 times, the base composition of
which were identical. Between the two repeat units,
there was a 99 bp spacer, which was an incomplete
copy of the two repeated units. In Asio otus also,
there were two repeated sequence units, a 127 bp se-
quence that was repeated eight times and a 78 bp se-
quence that was repeated six times, with a 55 bp
spacer. The control region of Athene noctua contains
three sets of repeats: an 89 bp sequence that was re-
peated three times, a 77 bp sequence that was re-
peated four times, and a 71 bp sequence that was re-
peated six times. The 89 bp unit represented an entire
77 bp unit plus an extra 12 bp unit to the 3′ end, with
only a 2 bp spacer between the two sets of repeat se-
quences. The 77 bp repeats and 71 bp repeats were
separated by 64 bp incomplete copies. Strix aluco,
however, had only one repeated sequence, a 78 bp
sequence that was repeated five times. The sequences
of the repeated units are shown in Table 4.
Low sequence homologies exist between the re-
peated units of the four owls and those of the other

Table 2 Nucleotide composition of mtDNA control region of the four species of Strigiformes
Species %A %C %G %T %A+T
Repeated sequences
%A+T
63.42
70.89
67.52
67.69
Asio flammeus
Asio otus
Athene noctua
Strix aluco
32.30
32.16
32.76
30.06
29.01
25.39
25.88
26.72
10.61
9.62
11.51
13.39
28.07
32.83
29.85
29.83
60.37
64.99
62.61
59.89


Fig. 1 General structure of mitochondrial DNA control region of the four species of Strigiformes
GLU = tRNAGlu; PHE = tRNAPhe; b = Bird similarity box. The number beneath the conserved sequence blocks denotes its length;
126*7 + 99 + 78*14 indicates a 99 bp interval between seven repeats of 126 bp and 14 repeats of 78 bp.

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Table 3 Comparison of CR conserved sequence block of the four species of Strigiformes
Note: 1 denotes Asio flammeus; 2 denotes Asio otus; 3 denotes Athene noctua; 4 denotes Strix aluco. * denotes identical base among the four species

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XIAO Bing et al.: Comparative Analysis of Complete Mitochondrial DNA Control Region of Four Species of Strigiformes 969


Table 4 Nucleotide sequences of repeated units in the control regions of the four species of Strigiformes
Species Repeated units Sequences
Asio flammeus



126 bp unit


78 bp unit
5′-TAACATCGGCGCCCAAATTCCGCTAACAATTTTGATCCCATCTATCCATTTTTCTAT
TCAACTTTCATCAAAACTTCTAGGCAAACTCTAACTGTTTGCCATTCGATCTTATCA
CTCACCTTACCC-3′
5′-TAGGCAAACTATAACAATCAATTATCGTCCACCATCAAAAATTTTACCGATCAATC
GAACGAACGATCAAAAACCTTC-3′

5′-TAGGCAAACTATAACTGTTTGCCAATCAACTTTATCGTTAATTCTTATCATAATATC
GGCACCCAAATCCCGCTAATAAATCTATCTTTTATCCATTTATTTTTTACCCAAATTTA
CTCGACACTTT -3′
5′-CAAACGAACGATAAGAAACTTTCTAGACAAACTATAACAATCAATTATCATCCAT
CATCAAAAATTTTATTGATCAAT-3′

5′-AAACTTATCCTCAATATAACCCTAAGTTTGCCTACAGAAATTAAACACGAATTCTA
TTATCCATTCTATTTTCCATTTATTTTAATCTT - 3′
5′-AAACTTACCCTAAATACAACCCTAAGTTTGCCTGCATAGATTAGGTATCGATCCTA
TCATCCATCCAAACTTTACAC-3′
5′-TAAAAGGTTGACCAAACAACCAAGCACTAATCAGAAATTCTTAACAAAAACCAA
TACTCATAATCGTAACT -3′

5′-AACAAACCATAGGGGAATTCTAGAGGAATTGCAACGATCATTTTGTATTCATCAC
CAAAAATTTTTATCGATCAATCA-3′

Asio otus




127 bp unit


78 bp unit

Athene noctua






89 bp unit

77 bp unit

71 bp unit


Strix aluco

78 bp unit

birds. Between Asio flammeus and Asio otus, the ho-
mology between the 126 bp (127 bp) unit and the 78
bp unit was also low. However, the corresponding
repeat units exhibited very high homology: the ho-
mology of the 78 bp unit was about 88.5% and that of
the 126 bp (127 bp) unit was about 75.8%. Moreover,
the homology of the 78 bp unit between Strix aluco
and Asio flammeus or Asio otus was about 70%. Be-
cause the 89 bp unit in Athene noctua was essentially
a 77 bp unit plus a 12 bp unit, it was compared to the
78 bp unit, and the homology was approximately
74%.
Potential secondary structures of the repeated
sequences in the control region were constructed us-
ing RNAstructure4.2 program (Fig. 2). Results
showed that this region was able to form a stable
stem-and-loop structure. MEME-Motif discovery tool
was used to analyze the eight repeated units. Results
seemed to indicate that there were many conserved
motifs in the repeated sequences, which are identified
in color in Table 5.
3 Discussion
3. 1 Basic characteristics of the control region
The size of the control region of avian mtDNA
is usually 1–1.5 kb. Few vertebrates have a control

Table 5 The conserved motif of repeated units in the CR of the four owls
Name
1
2
3
4
5
6
7
8
Start
78
1
1
24
6
6
10
21
P-value
6.73e-12
1.32e-16
1.09e-11
9.56E-17
3.44e-12
8.57e-12
3.00e-09
4.08e-12
Sites

Note: In the column Name, 1 and 2 denote the 126 bp and 78 bp repeated units of Asio flammeus, respectively; 3 and 4 denote the
127 bp and 78 bp repeated units of Asio otus, respectively; 5, 6, and 7 denote the 89 bp, 77 bp, and 71 bp repeated units of Athene
noctua, respectively; and 8 denotes the 78 bp repeated unit of Strix aluco. The number in the Start column denotes the site of the
repeated unit.

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Fig. 2 Secondary structure of repeat units of the four owls
A: the 78 bp repeat unit of Asio flammeus; B: the 126 bp repeat unit of Asio flammeus ; C: the 78 bp repeat unit of Asio otus; D: the
127 bp repeat unit of Asio otus ; E: the 78 bp repeat unit of Strix aluco ; F: the 89 bp repeat unit of Athene noctua ; G: the 77 bp
repeat unit of Athene noctua; H: the 71 bp repeat unit of Athene noctua.

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XIAO Bing et al.: Comparative Analysis of Complete Mitochondrial DNA Control Region of Four Species of Strigiformes 971


region that is longer than 3 kb. The longest reported
control region in vertebrates is that of Myxine gluti-
nosa[11] (3 628 bp, AJ404477). In this study, the
complete mtDNA control region of four species of
Strigiformes has been determined, and it was ob-
served that the length of the control region was un-
usually long, ranging from 1 771 bp to 3 290 bp. It is
noteworthy that Asio flammeus has the longest con-
trol region (3 290 bp) among all sequenced avian mi-
tochondrial DNA control regions, as shown in this
study.
The general organization of the control region of
the four species is similar to that reported for other
birds. The control region may be divided into three
domains, and some conserved sequence motifs could
be found in all the three domains. The tRNAGlu-
adjacent domain Ⅰ contains terminal-associated
sequences (TAS) and CSB-1-like conserved se-
quences. TAS is associated with termination of
H-strand replication, whereas the CSB-1-like motif is
considered to be unique to Aves [12]. B-box, C-box,
D-box, E-box, and F-box, present in domain Ⅱ in
most vertebrates, also exist in the four owls. They
may be associated with regulating H-strand synthesis.
In addition, some considered that B-box, D-box, and
F-box were prevalent in Aves but that C-box and
E-box existed only in some Aves[13], which implied
that B-box, D-box, and F-box might be more essential
in H-strand replication compared with C-box and
E-box. Bird similarity box could be also identified in
the four owls, and exhibited the highest similarity
among homologous sequences in domain
3). It suggests that bird similarity box may play a key
role in the replication and transcription of the mito-
chondrial genome in Aves. Furthermore, CSB-1 motif,
being located in the 5′ end of domain
served in the four owls. However, CSB-2 and CSB-3
motifs identified in mammals have not been found in
the four species [14]. Short conserved sequence blocks
(CSB-1, CSB-2, CSB-3) are usually considered to
regulate the initiation of replication and transcription
of the mitochondrial genome[15]. These above-men-
tioned results seem to indicate that CSB-1 plays a
(Table
Ⅱ
, is also co
Ⅲ
n-
more dominant role than does CSB-2 and CSB-3.
Furthermore, the four owls lacked heavy-strand rep-
lication origin OH and the bidirectional transcription
promoter LSP/HSP, which were extremely important
in heavy-strand replication and initiation of the bidi-
rectional transcription of mitochondrial genome in
most vertebrates. It is speculated that these four spe-
cies may form special secondary structures to carry
out these tasks.
3. 2 Origin and evolution of tandem duplication
sequences
The results of this study suggest that the large
size of the mitochondrial control region of the four
owls may be attributed to numerous repeated se-
quences within the 3′ end of their control regions.
Extensive tandem repeats in the mtDNA control re-
gion have been found in many vertebrate species (Ta-
ble 6). Nevertheless, such repeated sequences herein
may be a peculiar event. These repeat units in the CR
were compared among the four species and with other
vertebrates (Table 6). However, because of the lack of
data for mtDNA control region in other Strigiformes,
it is yet to be confirmed whether multiple repeated
sequences are a common feature in Strigiformes.
The results of this study also showed that very
high sequence homology existed in the 126 bp (127
bp) and 78 bp units in Asio flammeus, Asio otus, and
Strix aluco, which suggested that these corresponding
repeated units might have a common origin. They
might have existed before species divergence, al-
though independent evolution may have led to dif-
ference in these corresponding repeated units in the
four owls. In addition, because the 89 bp unit is a 77
bp unit with an extra 12 bp added to the 3′ end in
Athene noctua, the 89 bp and 77 bp units may also
originate from an ancestral sequence despite some
sequence differences.
Molecular mechanisms, such as recombination
and transposition[17], unequal crossing-over (gene
conversion)[18], and slipped-strand mispairing[19-21],
have been proposed on the origin of tandem repeated
sequences and subsequent generation of mtDNA

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Table 6 Comparison of the repeat units in CR between the four owls and the other vertebrates

Species Repeat unit in CR Derived source
This study
This study
This study
This study
Asio flammeus
Asio otus
Athene noctua
Strix aluco
126 bp ×7, 78 bp ×14
127 bp ×8, 78 bp ×7
89 bp ×3, 77 bp ×4, 71 bp ×6
78 bp ×5
27 bp ×2, 97 bp ×2, 11 bp ×4, 11 bp ×13 (+8 bp)
9 bp ×3, 11 bp ×2, 23 bp ×3
68 bp ×3, 56 bp ×5(+11 bp)
71 bp ×3(+43 bp)
53 bp ×2(8 bp spacer), 19 bp ×2
26 bp ×2(+43 bp), 4 bp ×18
56 bp ×5(+24 bp)
22 bp ×3, 21 bp ×14
10 bp ×4,10 bp ×11
6 bp ×54, 14 bp ×3
6 bp ×40, 8 bp ×11, 8 bp ×7
19 bp ×3, 9 bp ×12, 19 bp ×8
9 bp ×5, 11 bp ×2, 17 ×16
Buteo buteo
AF380305
Falco peregrinus
Ciconia ciconia
Gallus gallus
Rhea americana
Smithornis sharpei
Alligator mississippiensis
Diceros bicornis
Halichoerws grypus
Phoca vitulina
Didelphis virginiana
Ornithorhynchus anatinus
Note: The number in parentheses denotes one incomplete repeat.
NC000880
AB026818
NC001323
NC000846
NC000879
Y13113
L22010
X72004
X63726
Z29573
X83427

length variants. Although recombination is usually
thought of as the dominant mechanism that causes
variation in nuclear genomes, direct evidence for the
same is yet to be found in animal mitochondrial ge-
nomes. The results of this study seem to indicate that
the origin of tandem repeats of mtDNA control region
may result from slipped-strand mispairing during
mtDNA replication. It is proposed that the origin of a
new repeated sequence may be generated through the
following three steps. First, an extraneous and muta-
tional sequence should have been deleted from the
original sequence; second, the sequence loops out
because of strand contraction; and third, the sequence
expands to form a repeated sequence. As a result, the
extraneous sequence is not deleted but a new se-
quence is generated by slipped-strand mispairing.
Wilkinson and Chapman argued that because of uni-
directional replication of the H-strand in the con- trol
region, duplication of motifs should occur only in
one direction[22]. According to this hypothesis, repeats
at the 5′ end can be duplicated or deleted, whereas
repeats at the 3′ end cannot be duplicated and are
usually fragmentary due to partial deletion. In this
study, spacers of variable lengths have been found
between two repeat units (99 bp in Asio flammeus, 55
bp in Asio otus, and 64 bp in Athene noctua). They
are composed of incomplete copies of the 5′ end of
the first repeat unit and the 3′ end of the second re-
peat unit. This could be due to the duplication of the
two repeated sequences in opposite directions.
Therefore, according to Wilkinson-Chapman hy-
pothesis, the latter repeat sequences may be generated
by the replication of the L-strand.
The sequence polymorphism within the repeated
arrays results in an intermediate stage in the replica-
tion process, and its observation requires a balance
between two opposing evolutionary forces: point
mutations and insertion-deletion mutations [23]. The
former causes divergence, whereas the latter causes
homology. In this study, all 14 copies of the 78 bp
unit are exactly the same in Asio flammeus, which
may imply that insertion-deletion mutations play a
more dominant role than point mutations.
3. 3 Application of tandem duplication sequences
Several researchers believed that tandem re-
peated sequences are redundant in the mitochondrial
genome[24]. However, others hold opposing views [25].
In addition, palindromic and hairpin sequences lo-
cated within the repeated sequences have been re-
ported in many species and are capable of folding
into complex secondary structures[24]. The repeated

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XIAO Bing et al.: Comparative Analysis of Complete Mitochondrial DNA Control Region of Four Species of Strigiformes 973

sequences in the control region of the four owls are
able to form a stable stem-and-loop structure, which
is believed to be functionally essential in terminating
mitochondrial genome replication[12]. In addition,
there are many conserved motifs in the repeated se-
quences (Table 6), which may offer multiple copies of
functionally important sequences and even confer
replicative advantages to mtDNA molecules with
multiple copies of replication signals. In this regard,
repeated sequences may also be involved in the for-
mation of complex secondary structures of the control
region and strengthen the replication and transcription
of mtDNA. The results of this study showed that the
repeated sequences of the control region exhibited
some differences between species, even between two
individuals of the same species. It is therefore suggest
that the mtDNA control region may be used as a mo-
lecular marker to study population evolution.
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鸮形目 4 种鸟类线粒体调控区全序列的测定与比较研究
肖 冰1, 马 飞1, 孙 毅2, 李庆伟1
1. 辽宁师范大学生命科学学院，大连 116029；
2. 厦门大学生命科学学院，厦门 361005
摘摘 要要: 利用 Long-PCR 和 Primer Walking 的方法对鸮形目的短耳鸮、长耳鸮、纵纹腹小鸮、灰林鸮 4 种鸟类的线粒体调控
区进行了全序列测定。结果表明：短耳鸮的调控区长度为 3 290 bp；长耳鸮为 2 848 bp；纵纹腹小鸮为 2 444 bp；灰林鸮为
1 771 bp。短耳鸮的调控区长度是 4 种鸮中最大的，并且是目前已知最大的鸟类线粒体调控区。这 4 种鸮类调控区的基本
结构和其他鸟类相似，按照碱基变化速率的不同可以分为 3 个区: 碱基变化速率较快的外围区域Ⅰ、Ⅲ和保守的中间区域
Ⅱ。这 4 种鸟类调控区的 3′ 端均存在大量的串联重复序列，短耳鸮为 126 bp 单元重复 7 次和 78 bp 单元重复 14 次；长耳
鸮为 127 bp 单元重复 8 次和 78 bp 单元重复 6 次；纵纹腹小鸮有 3 个重复单元，分别为 89 bp 单元重复 3 次、77 bp 单元重
复 4 次和 71 bp 单元重复 6 次；灰林鸮仅有 1 个单元的串联重复为 78 bp 重复 5 次。调控区中串联重复序列可能是由链的
滑动错配产生，另外这些重复序列都能形成热力学稳定的多重茎环二级结构，而且在重复序列中还发现一些保守基序，这
说明重复序列可能具有一定的生理功能，影响调控区的调控功能从而影响线粒体基因组的复制和转录。
关键词关键词: 鸮形目；线粒体基因组；调控区；重复序列
作者简介作者简介: 肖冰（1980-） ，女，吉林人，硕士，研究方向：细胞生物学。E-mail: xiaobing19800516@sina.com