A Complete Sequence and Transcriptomic Analyses of
Date Palm (Phoenix dactylifera L.) Mitochondrial
Yongjun Fang1,2,3., Hao Wu1,2,3., Tongwu Zhang1,2,3., Meng Yang1,3, Yuxin Yin1,3, Linlin Pan1,3,
Xiaoguang Yu1,3, Xiaowei Zhang1,3*, Songnian Hu1,2,3*, Ibrahim S. Al-Mssallem1,3,4*, Jun Yu1,2,3*
1Joint Center for Genomics Research (JCGR), King Abdulaziz City for Science and Technology (KACST) and Chinese Academy of Sciences (CAS), Riyadh, Kingdom of Saudi
Arabia, 2James D. Watson Institute of Genome Sciences, Zhejiang University, Hangzhou, China, 3CAS Key Laboratory of Genome Sciences and Information, Beijing
Institute of Genomics (BIG), Chinese Academy of Sciences (CAS), Beijing, China, 4Department of Biotechnology, College of Agriculture and Food Sciences, King Faisal
University, Hofuf, Kingdom of Saudi Arabia
Based on next-generation sequencing data, we assembled the mitochondrial (mt) genome of date palm (Phoenix dactylifera
L.) into a circular molecule of 715,001 bp in length. The mt genome of P. dactylifera encodes 38 proteins, 30 tRNAs, and 3
ribosomal RNAs, which constitute a gene content of 6.5% (46,770 bp) over the full length. The rest, 93.5% of the genome
sequence, is comprised of cp (chloroplast)-derived (10.3% with respect to the whole genome length) and non-coding
sequences. In the non-coding regions, there are 0.33% tandem and 2.3% long repeats. Our transcriptomic data from eight
tissues (root, seed, bud, fruit, green leaf, yellow leaf, female flower, and male flower) showed higher gene expression levels
in male flower, root, bud, and female flower, as compared to four other tissues. We identified 120 potential SNPs among
three date palm cultivars (Khalas, Fahal, and Sukry), and successfully found seven SNPs in the coding sequences. A
phylogenetic analysis, based on 22 conserved genes of 15 representative plant mitochondria, showed that P. dactylifera
positions at the root of all sequenced monocot mt genomes. In addition, consistent with previous discoveries, there are
three co-transcribed gene clusters–18S-5S rRNA, rps3-rpl16 and nad3-rps12–in P. dactylifera, which are highly conserved
among all known mitochondrial genomes of angiosperms.
Citation: Fang Y, Wu H, Zhang T, Yang M, Yin Y, et al. (2012) A Complete Sequence and Transcriptomic Analyses of Date Palm (Phoenix dactylifera L.)
Mitochondrial Genome. PLoS ONE 7(5): e37164. doi:10.1371/journal.pone.0037164
Editor: Bernd Schierwater, University of Veterinary Medicine Hanover, Germany
Received October 31, 2011; Accepted April 16, 2012; Published May 24, 2012
Copyright: ? 2012 Fang et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: The project is supported by a project grant from King Abdulaziz City for Science and Technology, Riyadh, Saudi Arabia [428-29]. The funder had no role
in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
* E-mail: firstname.lastname@example.org (JY); email@example.com (XZ); firstname.lastname@example.org (SH); email@example.com (ISAM)
. These authors contributed equally to this work.
The widely-accepted hypothesis about the origin of the
mitochondrion assumes that it descended from an endosymbiontic
event involving an a-proteobacterium-like organism and the
common ancestor of eukaryotes . Evolving from algae to land
plants, including bryophytes and angiosperms, plant mitochondri-
al (mt) genomes have increased their sizes, especially in the non-
coding region. Among land plants, bryophytes, i.e., liverworts,
mosses, and hornworts, represent the basal forms. They have
similar gene order, genome size, and a fraction of non-coding
sequences [2,3]. As evolution continues, land plants gained new
mechanisms to facilitate frequent gene exchange between mito-
chondrial and chloroplast genomes as well as between mitochon-
drial and nuclear genomes [4,5]. For instance, mitochondrial
genomes of angiosperms have long been known for their slow
evolutionary rate , existence of subgenomic circles in addition
to a master genomic circle , extraordinarily large and highly
variable genome sizes , trans-splicing of group II introns ,
high density of RNA editing [10,11], divergent non-coding
sequences , and frequent gene transfer . The inter-
genomic gene transfer, together with the continuing increase of
non-coding DNA sequences, leads to a broad size range in
angiosperm mt genomes, which as of today is from ,200 to
2400 kb based on the known mt sequences and experimental
estimations [8,14]; up to date (July, 2011), there have been more
than 40 plant mt genomes sequenced, including 22 angiosperm mt
Phoenix dactylifera L., also known as date palm, is economically
the most important plant in the Middle East and North Africa
, and it is estimated to have more than 450 cultivars or
varieties in the Kingdom of Saudi Arabia and nearly 2,000
varieties around the world . Therefore, sequencing its
mitochondrial genome, together with its nuclear  and
chloroplast genomes , is of essence in improving its
agricultural, horticultural, and nutritional values. In this study,
combining data from two next-generation sequence platforms,
pyrosequencing (Roche GS FLX) and ligation-based sequencing
(Life Technologies SOLiD), we assembled P. dactylifera mt-genome
(cultivar Khalas, Al-Hasa Oasis, Saudi Arabia) –the first from the
Arecaceae family. In addition, analysis of the mt genome sequence
PLoS ONE | www.plosone.org1May 2012 | Volume 7 | Issue 5 | e37164
and transcriptomic data are of importance in revealing mechan-
isms underlying mitochondrial genome evolution and the unique
evolutionary status of P. dactylifera among angiosperms. Further-
more, based on the data from three commonly-grown cultivars, we
also investigated RNA editing sites and SNPs within the species.
Results and Discussions
General Features of P. dactylifera L. mt Genome
We assembled the P. dactylifera master mt chromosome into
a 715,001 bp circular molecule (Figure 1; the assembling details
are described in Materials and Methods) with an average GC
content of 45.1%; it is now the fourth largest mt genome
sequenced after those of Cucumis sativus (1,555,935 bp) ,
Cucurbita pepo (982,833 bp), and Vitis vinifera (773,279 bp). Its
protein coding sequence is composed of only 6.5% of the genome
(46,770 bp) and this gene content is similar to other published
angiosperm mt-genomes (Table S1 and Table S2). The rest, also
the majority (93.5%) of the genome, is composed of non-coding
(the cp-derived regions are also considered as non-coding in this
regard), which harbors 0.33% tandem and 2.3% long repeats (the
repeat lengths are greater than 50 bp). RNA genes and intron
sequences are 1.1% and 4.3% of the mt genome, respectively. This
mt genome also contains the second highest proportion (10.3%) of
cp-derived sequences among the sequenced mt genomes to date, of
which several intact genes, such as petA, petG, petL, psaJ, psbT, rpl20,
rpl33, and rps8 are identified. Since the age of the cp-derived
sequences or time when the sequences inserted into mt genomes
varies greatly , we are unable to prove whether these genes are
actually transcribed or active since we extracted the total RNA
(contains both nuclear and organellar transcripts) from each tissue
for constructing transcriptomic libraries among which the
expression data of the cp-derived sequences and authentic cp
sequences are impossible to separate (see Materials and Methods
for more details). Most of P. dactylifera mt genome is rather
diverged from other angiosperms. For example, only ,21% of P.
dactylifera mt genome sequence is shared (over 70% identity) by
Vitis, Oryza and Bambusa, and even less by Zea (,15%) and
Arabidopsis (,11%). In addition, consistent with results from
previous studies, we observed that three co-transcribed gene
clusters, 18S-5S rRNA, rps3-rpl16, and nad3-rps12, are conserved in
other angiosperm mt genomes . We summarized general mt
genome features including size variations, AT content, and intron
types of 15 non-redundant sequenced plant mt genomes (including
12 higher plants and three lower plants) in Table 1. Our
phylogenetic analysis based on 22 concatenated conserved genes
among 15 selected mt genomes (Figure 2) revealed that P. dactylifera
appears to be the more basal among monocots.
Protein Coding, rRNA, and tRNA Genes
The P. dactylifera mt genome contains at least 38 protein-coding
genes and five complete ORFs, most of these genes encode
proteins of the electron transport chain, such as nine subunits of
nicotinamide adenine dinucleotide dehydrogenase (complex I),
apocytochrome b (complex III), three subunits of cytochrome c
oxidase (complex IV), five subunits of ATP synthase F1 (complex
V) and four cytoplasmic membrane proteins required for
cytochrome c maturation (Table 2).
We compared these protein coding genes to 11 other sequenced
angiosperm mt genomes (Table S2). First, P. dactylifera mt genome
does not have the genes encoding respiratory chain complex II,
such as sdh3 and sdh4, which are only found in two dicots, Nicotiana
tabacum and V. vinifera. Second, our assembly is similar to V. vinifera,
and both contain one copy of RNA polymerase gene harboring
a conserved domain characteristic of pfam00940 superfamily of
polymerases . Third, rps14 present in Brassica napus and V.
vinifera is also found here, whereas rps11, another ribosomal protein
gene, is exclusively detected in our assembly. Both genes have full
open reading frame (ORF) and are likely functional in date palm,
though in many other known angiosperm mt genomes they are
either pesudogenes or transferred into nuclear genomes [23,24].
Fourth, the recently identified rpl10 gene, being identified as orf-
bryo1 in vascular plants and charophycean green algae  and
orf168-related sequences in bryophytes and angiosperms ,
seems to be interrupted in our assembly and possibly because of
a frame shift event. Fifth, we found several pseudogenes in our
assembly, which appear intact in other mt genomes, such as orf99-
b (as orf100-y in our gene list) in Zea mays and cp-derived gene
psbT in V. vinifera. In addition, some of the universally expressed
ribosomal genes, including three rRNA genes (5S, 18S, and 26S
ribosomal RNA genes), are also unambiguously identified ; 5S
and 18S rRNA genes are also closely related and distant from 26S
rRNA gene in date palm mt genome.
A genome-wide screening
sequences (Table S3) in our assembly; among them, 12 seem to
be cp-derived, which exhibit higher sequence identity (.98%) to
their chloroplast counterparts than their mitochondrial counter-
parts , and three predicted tRNAs seem to have introns.
Among these 30 tRNA genes five amino acids (A, L, R, T, and V)
are not encoded, although tRNAs for 20 amino acids are necessary
for protein synthesis in mitochondria. In addition, having
compared the date palm tRNA gene content to those of seven
other plants mt and cp genomes (Figure 3), we conclude that there
are 10 tRNA genes, among which nine encoding tRNAs for the
five amino acids, are actually lost after the divergence of liverworts
from seed plants. These results suggest that the missing tRNAs are
supplied by either the chloroplast or nuclear genomes. In addition,
we found that four mt tRNA genes of higher plants are gradually
lost and replaced by cp-derived tRNA . The reason why mt
tRNA genes are replaced by both cp-derived and nuclear
counterparts remains an open question. There is also another
possibility–all mt tRNA genes may eventually be replaced and
what we observed here is only an intermediate and dynamic
Plastid DNA Insertions
Chloroplast and mitochondrial genomes are known to share
sequences due to frequent gene transfer events [5,29,30]. Frequent
DNA transfer from cpDNA to mtDNA occur as far back as the
common ancestor of the extant gymnosperms and angiosperms,
about 300 MYA (million-years-ago) . Our assembly contains
more than 100 fragments of chloroplast origin (over 80% identity),
ranging from 50 to 6,521 bp in length (Table S4). The total
fraction of chloroplast DNA sequences present in P. dactylifera mt
genome is 73,691 bp, corresponding to 10.3% of the whole mt
genome, and 46.5% of P. dactylifera cp-genome. The proportion of
cp-derived sequences in our mt genome assembly is comparable to
the two large sequenced plant mt genomes, V. vinifera (8.8%) and
Cucurbita pepo (11.6%) , but larger than those of the other
known plant mt genomes (Table 1). These results suggest that
chloroplast DNA sequence insertion is an important mechanism
for plant mitochondrial genome size expansion and sequence
Most of chloroplast sequence insertions in P. dactylifera mt
genome are unique, as evident from the observation that only
nine out of 44 insertions (over 200 bp) have full-length
homologous sequences shared by other known mt genomes
(.90% length coverage, .70% identity) (Table 3). Among the
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org2 May 2012 | Volume 7 | Issue 5 | e37164
nine cp-derived homologous sequences, six and five of them are
also found in Vitis and Bambusa, respectively, whereas none is
found in A. thaliana. These nine insertions tend to have higher
GC content, resembling that of mt genomes as compared to the
unique and possibly new insertions (Figure S1), which suggests
that these cp-derived sequences did, in some extent, gradually
increase their GC content to become similar to their host
Introns and RNA Editing
We identified 23 group II introns in 10 protein-coding genes,
including four trans-spliced introns of nad1 and nad5, and 20 cis-
spliced introns in ccmFc, cox2, nad1, nad2, nad4, nad5, nad7, rpl2, and
rps3. No group I intron was discovered in our assembly. In general,
the functional mitochondrial rRNA and tRNA genes of the
sequenced angiosperm mt genomes do not possess introns, but we
found three intron-containing tRNA genes in our assembly: trnK-
TTT, trnN-ATT and trnSup-CTA, and we have yet to validate if
they are functional or not.
Mitochondrial RNA editing is essential for functional protein
synthesis since nearly all plant mt mRNAs are edited [32,33,34]
and it modifies amino acids and generates new start or stop codons
[35,36,37,38,39], and it has been documented in most plants
except algae and mosses. It suggests that this cellular process is
ancient arisen in early land plants after they split from Bryophyta
. We predicted nearly 600 putative RNA editing sites (Table
S5) using PREP-Mt –an effective tool identifying C-U editing
sites. We found that the nad4 gene contains the most edited sites
(59). In addition, our comparative analysis revealed that 305
(51.5%) and 278 (47.0%) C-U editing sites in date palm are shared
by O. sativa and A. thaliana, respectively (Figure 4). Experimental
examination confirmed 40 of 41 predicted C-U editing sites in five
randomly chosen genes (atp1, atp4, atp9, rpl116 and rps19) using
cDNA sequences (Table S6) and additional nine sites not detected
Figure 1. A circular display of P. dactylifera mitochondrial genome. We display (starting from outside to inside): physical map scaled in kb,
coding sequences transcribed in the clockwise (red) and counterclockwise directions (blue), chloroplast-derived regions (green boxes), sequence
repeats (black), histogram of transcriptome data (green bar, standing for average RPKM value per 200 bp, transformed using natural logs and ranging
from 0 to 10), GC content variations (brown bar in a 500 bp sliding window and 500 bp increments), and SOLiD mate-pair (MP) read validation
(sliding window 2 kb, MP insertion size 5–6 kb, Step size 15 kb). This figure was generated by using the Circos program . Y indicates pseudogene.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org3 May 2012 | Volume 7 | Issue 5 | e37164
Table 1. Comparative analysis of genomic features among 15 mt genomes.
Gene number (Total/Protein/tRNA/rRNA)
Group II introns (Cis/
We summarized several genomic features from 15 representative mt genomes, including AT content of the mt genomes, the percentage of gene-coding sequences, and the percentage of chloroplast-derived sequences in mt
genome sequences. We only used the genus names for the reference genomes.
aInformation about these mt genomes are from reference  and information about other plant mt genomes are either from original publications or NCBI databases (see Table S1).
bTo be consistent, repetitive sequence contents in the 15 plant mt genomes are all computed by using REPuter (length .50 bp; mismatch #3).
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org4 May 2012 | Volume 7 | Issue 5 | e37164
by PREP-Mt were identified. We also compared their tissue
disparity between mRNA transcripts extracted from yellow and
green leaves, but no obvious tissue-specific RNA editing patterns
are yet identified among these five genes, although reports in the
literature indicated that the extent of atp6 editing is significantly
different among tissue types . Therefore we assume the tissue-
Figure 2. Phylogeny inferred from 22 genes common to 15 plant mt genomes. We constructed an ML tree using PHYML (version 3.0) 
(Chara vulgaris as outgroup, see Materials and Methods for details). Nodes receive over 90% bootstrap replicates are indicated. P. dactylifera mt
genome rooted at the basal position of monocots (red).
Table 2. The gene content of P. dactylifera mt genome.
Genes of Mitochondrial Origin
Complex I nad1, nad2, nad3, nad4, nad4L, nad5, nad6, nad7, and nad9
Complex III cob
Complex IVcox1,cox2, and cox3
Complex V atp1, atp4, atp6, atp8, and atp9
Cytochrome c biogenesis ccmB, ccmC, ccmFc, and ccmFn
Ribosome large subunitrpl2, rpl5, and rpl16
Ribosome small subunit rps1, rps2, rps3, rps4, rps7, rps11, rps12, rps13, rps14, and rps19
Intron maturase matR
SecY-independent transporter mttB
rRNA genes 5sRNA,18sRNA, and 26sRNA
tRNA genes trnC-GCA, trnD-GTC, trnE-TTC, trnF-GAA, trnK-TTT(62), trnM-CAT(64), trnN-ATT, trnP-TGG(62), trnQ-TTG, trnS-GCT,
trnS-GGA, trnS-TGA, trnSup-CTA, trnT-TGT, and trnY-GTA
Hypothetical genes5 ORFs
Genes of Chloroplast Origin
Genes with intact ORFsa
accD, atpI, cemA, infA, matK, ndhI, ndhJ, petA, petB, petG, petL, psaB, psbA, psbE, psbH, psbJ, psbL, psbN, psbZ, rpl14,
rpl33, rpl36, rpoA, rps14, rps2, rps4, and ycf4
PseudogenespsbT and rps18
tRNA genes trnC-GCA, trnF-GAA, trnG-GCC, trnH-GTG, trnM-CAT, trnN-GTT, trnP-GGG,trnS-TGA, and trnW-CCA(62)
Genes of Nuclear Origin
aGenes with intact ORFs in cp-derived regions are identified based on .95% identity and .95% length coverage to the known cp genes.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org5 May 2012 | Volume 7 | Issue 5 | e37164
specific RNA editing patterns may be only detectable in certain
genes, cell types, and developmental stages .
Plant cells usually possess hundreds to thousands mitochondria
or copies of mt genomes that can be regarded as a population
when genetic heterogeneity is to be investigated. High throughput
next-generation sequence technologies provide us the opportunity
to survey single nucleotide polymorphisms in the same or different
species (subspecies or cultivars) by mapping reads to a reference
sequence and to each cultivar. The polymorphisms within the
same cultivar genome (intra-varietal SNPs) and among different
cultivars genomes (inter-varietal SNPs), discovered by high-
coverage of reads, can also be separated into major and minor
genotypes based on simple read counts. Here, we use three runs of
SOLiD fragment data from each of the three cultivars (Khalas,
Fahal, and Sukry) sequenced in our study for intra-varietal
(Table 4) and inter-varietal SNP identification (Table 5). We
identified 651, 703, and 731 intra-varietal SNPs in cultivar Khalas,
Fahal, and Sukry, respectively, estimated to have a polymorphism
rate of one in 1,000 bp, which is about two times higher than that
of date palm chloroplast  but is only about one tenth of rice mt
genome . We should be cautious here since different SNP
analysis methods are applied because of the distinct sequencing
strategies used in sequencing these mt or cp genomes. The rates of
each variation type among these intra-varietal SNPs of the three
cultivars are very similar except the types (such as A to T or G to C
and vice versa) that do not change GC contents are less
represented. These SNPs can also be separated into transition
and transversion types, and as a result, there are 297, 325, 287
transitions and 354, 378, 347 transversions for Khalas, Fahal, and
Sukry, respectively. The rate of transversions is slightly higher than
Figure 3. The distribution of tRNAs in vascular and angiosperm plant mitochondrial and chloroplast genomes. Native tRNA genes in
mitochondrial and chloroplast genomes are shown in open square and circles, respectively. Solid squares indicate cp-derived tRNAs found in mt
genome and y stands for pseudogene. There are ten tRNAs (their anticodons are highlighted in bold) that are gradually lost in genome evolution and
four tRNAs (their anticodons are underlined) that are gradually replaced by their cp-derived counterparts. These eight mt genomes are listed
according to their relative phylogenetic positions in Figure 2.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org6 May 2012 | Volume 7 | Issue 5 | e37164
that of transitions, though in chloroplast transversion (52) is 26
that of transition (26) .
All together, there are 120 candidate SNP sites identified among
the three cultivars (Table 5), with an inter-varietal polymorphism
rate of 0.017%, similar to that of subspecific (between subspecies)
polymorphisms between rice cultivar 93-11 and PA64S, ,0.02%
. The inter-varietal SNPs are predominantly found in non-
coding regions, only seven SNPs were found in coding sequences
(all are located in 26S and 18S rRNA genes; Table S7): two
between Khalas and Fahal, six between Khalas and Sukry, and six
between Fahal and Sukry (Table 5). As to the remaining 113 inter-
varietal SNPs residing in non-coding regions (Table S8), 79 of
them are between Khalas and Fahal, 91 between Khalas and
Sukry, and 50 between Fahal and Sukry (Table 5). Such
a distribution implies that Fahal and Sukry are more related than
either one of them to Khalas.
P. dactylifera mt genome has much less repetitive sequences as
compared to those of other known angiosperms . Only one
long palindromic sequence with repeat unit longer than 1000 bp
Table 3. The distribution of nine P.dactylifera chloroplast-derived mt regions in five known plant mt-genomes.
Arabidopsis VitisBambusa OryzaZea
87871–88837 96788 0.4224
179701–180523 82391 0.4702
535882–536375494 94 0.4231
271397–271847 45187 0.3792
586885–587154270 95 0.4870
We selected homologous sequences with identity .70% and length coverage .90% for the comparative analysis. The results for two dicots (Arabidopsis and Vitis) and
three monocots (Bambusa, Oryza, and Zea) are listed here. The presence (+) and absence (2) of the corresponding cp regions are indicated based on identity and length
coverage. Only the genus names are used for the reference mt genomes.
aThe sequence identity between the cp sequence insertions in P. dactylifera mt genome and their cp homologs.
bThe GC content of the cp-derived sequences in P. dactylifera mt genome.
Figure 4. Venn diagram of shared RNA editing sites among
three plant mt genomes.
Table 4. Intra-varietal SNPs among the three cultivars.
T/C66 66 64
C/A 6060 60
C/G11 14 10
G/C 10 1212
G/T 65 6363
T/G 7175 62
aMajor and minor genotypes are separated with oblique lines (/).
bNumbers of sites are calculated for each cultivar.
Table 5. Inter-varietal SNPs.
Khalas vs. Fahal2 79 81
Khalas vs. Sukry69197
Fahal vs. Sukry65056
Khalas vs. Fahal vs. Sukry7113120
a‘‘Coding’’ and ‘‘Non-coding’’ indicate numbers of inter-varietal SNPs found
among the groups.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org7 May 2012 | Volume 7 | Issue 5 | e37164
was identified (Table S9) and no inverted repeats were found.
Overall, long repeats (.50 bp) only account for 2.3% of the
genome, even lower than that of Vitis (2.9%) and Vigna radiata
(2.7%) , which contain the lowest repeat contents among
sequenced angiosperm mt genomes, whereas Tripsacum and Oryza
contains 36.4% and 30.4% long repetitive sequences, respectively.
This situation also applies to tandem repeats, which constitute only
0.33% of the genome (Table S10). Among the examined 15 plant
mt genomes, whose tandem repeat contents range from 0.08%
(N. tabacum) to 6.13% (Cycas taitungensis), only three mt genomes,
those from N. tabacum, O. sativa and Chara vulgaris, contain tandem
repeats lower than date palm (Figure 5). It is well known that plant
mitochondria are exceptionally flexible in genome size and
structure, and the accumulation of repetitive sequences often
results in high sequence divergence. For instance, Cucurbita mt
genome contains 38% of short repeats (19–621 bp in length) that
make it the largest reported mt genome so far , whereas maize
expanded its mt genome size by duplication of large sequences
. Therefore, it is rather unusual that date palm mitochondrial
genome is both lower in tandem repeat content and rare in large
duplications. It seems that larger mt genomes of angiosperms tend
to have shorter repeat lengths when long repeats are compared.
For instance, mt genomes of Cucurbita (982 kb), Vitis (773 kb), and
P. dactylifera (715 kb) have their largest repeat lengths of 621 bp,
651 bp, and 1,171 bp, respectively.
The mt-genome is transcribed by a phage-type RNA poly-
merase encoded in the nuclear genome . The transcription
process is rather complex characterized by splicing, editing,
terminus processing, and multiple promoters . In addition,
mitochondrial genome transcription is reported to be capable of
adapting to specific regulation . Here, in order to better
understand tissue-specific mt gene regulation and the contribution
of mt genes to the development of different tissues, we performed
a thorough transcriptome survey across eight date palm tissues
(Figure 1 and Figure S2) using high-performance next-generation
sequencers. We discovered that ,30.8% regions of our assembly
are transcribed (Table 6), slightly lower than that of the rice
(,48.5%) , with an average sequence coverage of ,446
calculated based on 40 conserved house-keeping genes (Figure 6).
On the one hand, our whole genome level gene expression
profiling indicated that two tissues, green leaf and fruit, have the
most abundant transcripts (Figure S2) but have the lowest gene
expression level in terms of RPKM value (reads per kilobase of
exon model per million mapped reads)  (Figure 6). Male and
female flowers, root, and bud, on the other hand, tend to have
higher gene expression levels but less in transcript abundance than
the leaves. We assume that developing tissues, such as yellow leaf,
bud, and root, need more energy than the relatively mature tissues,
such as green leaf and fruit. By the same token, the highly
expressed genes in female and male flowers are possibly related to
flower development that not only depends on a set of nuclear
genes but also on the coordinated action of mitochondrial genes
. It is possible that the variable expression of mt genome-
encoded genes is relevant to the copy number variation of mt
genomes (similar to the number of mitochondria per cell) [53,54]
or its changing status in tissue development. In addition, several
other obvious tissue-specific gene expression patterns can be
observed. First, consistent with a previous study that atp1 gene
prefers to express in pollen mother cells , we also detected that
the transcript of atp1 is obviously more abundant in male flower
than in other tissues examined. There is another gene matR that
encodes a maturase-related protein also expressed in a relative
higher level. Previous study revealed that this gene suffers from
modest RNA editing in maize and soybean and was predicted to
be functional . Our results here indicate that this gene should
have utmost importance in male flower development. Second, the
maturation process of yellow to green leaves seems to involve the
suppression of about half of the 40 mt house-keeping genes, and
the fact is further confirmed that developing tissues are more
affected by mt gene expression. Third, during seed maturation,
half of the genes were found to be up-regulated when compared to
the mt gene expression pattern in fruit. Fourth, interestingly, we
found that the gene expression patterns of yellow leaves and seeds
are quite distinct–down-regulated genes in one tissue tend to be
Figure 5. Percentage of long repeats and tandem repeats of 15 mt genomes. We analyzed long repeats (repeat unit .50 bp) using REPuter
 and tandem repeats based on Tandem Repeat Finder  (see Materials and Methods for details). The genus names are used to represent the
sequenced mitochondrial genomes and arranged according to their relative phylogenetic positions in Figure 2.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org8 May 2012 | Volume 7 | Issue 5 | e37164
highly regulated in the other tissue–except that of ccmFn, cox2, rps1,
rps3, and rps19 which have no obvious differences between these
two tissues. Fifth, two genes, rps1 and rps19, are found clearly
highly expressed in root as compared to other tissues. The
functional roles they play in root development still need further
experimental confirmation. Sixth, consistent with previous studies,
rRNA gene transcripts are found to be more abundant, ,9–13
fold than protein coding genes , but our large-scale
transcriptomic analysis reveals a much higher transcription level
changes ,50–400 fold than the average level of protein-coding
genes according to RPKM values, and the order of expression
levels for the ribosomal RNAs is 5S rRNA .26S rRNA .18S
As the first of the palm family plants, P. dactylifera mt genome
displays several unique features. First, it positions at the root of the
known monocot mt genomes. Second, it has a very low level of
repeat content and shows abundant RNA editing events. Third, it
exhibits a high level of chloroplast sequence insertions as
compared to other known angiosperm mt genomes. Furthermore,
our large-scale transcriptome analysis revealed that ,30.8% of its
sequences are transcribed and show obvious tissue-specific gene
regulation patterns, among which both female and male flowers,
root, and bud exhibit higher gene expressions than other sampled
tissues. Our complete mt genome sequence assembly represents
a new addition to the growing number of plant mt genomes in the
public databases and paves a way for further investigations on
mitochondrial biology of seed plants.
Materials and Methods
We used three domestic P. dactylifera cultivars, Khalas (male and
female), Sukry (female), and Fahal (male), for this study. Tissue
samples from adult date palm trees grown in Al-Hasa Oasis of
Kingdom of Saudi Arabia are harvested, including soft bud, flower
(male and female), fruit, root, yellow leaf (young), and green leaf
(old). We disinfected the samples with 75% ethanol and froze them
in liquid nitrogen immediately. For longer term storage, they are
stored in 280uC freezer until use.
Genomic and RNA Sequencing
The P. dactylifera mt genome sequences are produced as part of
the Date Palm Genome Project (DPGP, a joint effort between
KACST and CAS). Genomic DNA was extracted from 50 g soft
bud tissues according to the CTAB-based method. We used 5 mg
purified DNA for shearing and constructing fragment libraries
following the GS FLX Titanium general library preparation
protocol. The ssDNA libraries were amplified with emulsion-PCR
and enriched, and the samples were sequenced on Roche/454 GS
SOLiD long mate pair (LMP) libraries of the three cultivars
were constructed by following SOLiD Library Preparation Guide
(SOLiD 4.0) and at least 20 mg genomic DNA was used depending
on different insert sizes (600–6000 bp). After emulsion PCR and
beads enrichment (EZ beads system, AB), template beads of each
LMP library were deposited to 2 quarter of slide and then loaded
onto a SOLiD 4.0 instrument.
For transcriptomic study, tissue samples were grinded into fine
powder followed by CTAB-based RNA extraction, and 2.5 M
LiCl was used to remove polysaccharides. 0.5 mg rRNA-depleted
total RNA (RiboMinus Plant Kit, Invitrogen) were used to
construct transcriptomic libraries according to the instruction from
SOLiD Total RNA-Seq Kit.
Sequence Assembly and Validation
We separated candidate mt genome reads from eight Roche/
454 GS FLX runs based on 40 published plant mt genome
sequences (identity $80% and E-value #1025). About 1.5 millions
reads were obtained and assembled by using Newbler (version 2.3
with default parameters)–a de novo sequence assembly software
provided by Roche. As a result, we obtained 29 mt genome
contigs (total ,438 kb) with an average length of 15 kb. These
contigs were extended to 662 kb by adding additional Roche/454
reads. Subsequently, SOLiD mate-pair data (2650 bp libraries)
with insertion sizes of 1–2 kb and 3–4 kb were used to construct
scaffold (50-nt overlap cutoff and less than 2-nt mismatch). A total
of 3,918 homopolymers with repeat unit ranging from 5 to 11 were
verified and revised based on SOLiD fragment data using BFAST
program (version 0.6.4d) . At last, 715,001 bp complete mt
genome was assembled with an average sequence depth 1306.
The final genome sequence was validated by SOLiD LMP data
with insertion sizes of 3–4 kb, 4–5 kb, and 5–6 kb in a 2 kb sliding
window with variable step sizes; we show the result from an
analysis using 5–6 kb insert size in 15-kb step size in Figure 1.
The complete sequence of the date palm mt genome was
deposited to GenBank (accession number JN375330).
A preliminary annotation was carried out by mapping final
genome sequence with BLAST (identity .90% and over-
lap.90%) [59,60] hits to known mitochondrial genes, and
subsequently by testing for consistency of the ORFs using NCBI
online tool the ORF finder (http://www.ncbi.nlm.nih.gov/
projects/gorf/, the standard genetic code was applied). The exact
gene and exon boundaries were determined by alignment of
homologous genes from several common mt genomes (Table S2)
and verified based on transcriptomic data. The tRNA genes were
identified by using a local chloroplast and mitochondrial tRNA
database, BLAST search tools [59,60], and the help of tRNAscan-
SE program (version 1.4 and default parameters were used) .
Both group I and group II introns were predicted by using an
online software Rfam (version 10.1; http://rfam.sanger.ac.uk/;
default parameters) . Homology search using BLAST [59,60]
was carried out to identify chloroplast-derived regions in the mt
genome assembly (over 80% sequence identity; E value #1e-5;
length .50 bp).
Table 6. The transcript coverage of P.dactylifera mt genome.
=0 494769 69.20
aRead number in each genome position. ‘‘=0’’ means no transcription activity
was observed and the larger the number the higher the gene expression level.
Average coverage of 40 highly-conserved genes (,51,000 bp in length) in
P. dactylifera mt genome is 44.266.
bTotal length of genomic sequences defined for transcript expression level.
cThe proportion of transcribed region relative to the whole mt genome.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org9 May 2012 | Volume 7 | Issue 5 | e37164
RNA Editing Analyses
We predicted putative RNA editing sites in protein-coding
genes using the PREP-mt web-based program (http://prep.unl.
edu/) . To achieve a balanced tradeoff between the number of
false positive and false negative sites, the cutoff score (C-value) was
set to 0.6 as suggested by the author. All other parameters are set
to default values. We also verified some of the RNA editing sites in
Figure 6. Gene expression profiles of P. dactylifera mitochondrion among 8 tissues. We used 40 house-keeping (conserved over diverse
plant lineages) genes for hierarchical clustering (Manhattan distance method). Red and green indicate high and low levels of gene expression,
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org 10May 2012 | Volume 7 | Issue 5 | e37164
five genes (atp1, atp4, atp9, rpl116, and rps19) across the two leaf
tissues (yellow and green leaves) using cDNA data from Roche/
454 GS FLX system (NCBI accession number SRA045434.3).
The five genes are chosen randomly, whose cDNA sequences are
full-length and better in quality.
We carried out both intra-varietal and inter-varietal SNP
analysis across three cultivars: Khalas, Fahal, and Sukry. Three
runs of SOLiD LMP reads for each cultivar (about 60 Gb) were
mapped to the reference mt genome (Khalas) by using BioScope
software (version 1.3). The mapping results were then used for
SNP identification based on a Bayesian algorithm according to the
BioScope Software User Guide.
Analysis of Repetitive Sequences
ic, reverse, and complemented repeats, using the REPuter (version
2.74; with a minimal length of 50 bp and 3 mismatches) . We
tandem repeats using a tandem repeat finder (http://tandem.bu.
edu/trf/trf.html; default parameters wereused) .
We used 22 protein-coding genes (atp1, atp4, atp6, atp8, atp9,
ccmC, ccmFn, cob, cox1, cox2, cox3, nad1, nad2, nad3, nad4, nad4L,
nad5, nad6, nad9, rps3, rps4, and rps12) common to 15 plant mt
genomes for our phylogenetic analysis. We aligned the sequences
using Clustalw2 (default parameters were used) , removed
ambiguously aligned regions based on Gblocks (version 0.91b;
minimum number of sequences for a conserved position and
flanking position is set to 10 and 15, respectively; no more than
eight contiguous non-conserved positions and no gap are allowed)
, and concatenated the sequences. The maximum-likelihood
tree was constructed by using PHYML (version 3.0)  under
HKY85+C4 model (C. vulgaris is used as outgroup). The bootstrap
value was set to 100. All other parameters are set as default.
We used transcriptome data from bud, root, seed, fruit, male
and female flowers, yellow and green leaves of cultivar Khalas. On
average, ,700,000 SOLiD reads (50 bp with 3 mismatches or less)
are used from the libraries. RPKM values are measured (reads per
kilobase of exon model per million mapped reads)  and used to
estimate gene expression, which are calculated according to:
chloroplast-derived sequences. We defined chloroplast-de-
GC content variations between new and old
rived sequences unique to P. dactylifera mitochondrial genome as
‘‘New’’ and those shared by other plant mt genomes as ‘‘Old’’.
FF, female flower (,422,000 reads); MF, male flower (,589,000
reads); F, fruit (,1,048,000 reads); S, seed (,179,000 reads); B,
bud (,457,000 reads); GL: green leaf (,2,388,000 reads); YL,
yellow leaf (,606,000 reads); R, root (,545,000 reads); P, genes
on the positive strand; N, genes on the negative stand; and CP,
chloroplast-derived regions. Their RPKM values (transformed
using log10) range from 0 to 9 for genes on the positive strand and
0 to 29 for genes on the negative strand.
Transcriptome analysis across eight tissues.
15 plant mt genomes used in this study.
Genes in 12 angiosperm mt genomes.
tRNA gene content of P. dactylifera mt
fera mt genome.
Chloroplast-derived sequences in P. dactyli-
RNA editing sites in three different plant mt
tissues based on GS FLX reads.
RNA editing validation of five genes in two
the three cultivars.
Inter-varietal SNPs in coding regions among
among the three cultivars.
Inter-varietal SNPs in non-coding regions
dactylifera mt genome.
Long repeats (repeat unit . .50 bp) in P.
Tandem repeats in P. dactylifera mt genome.
We thank Jiucheng Liu, Dawei Huang, Guiming Liu, Quanzheng Yun,
Sun Zhang, Guangyu Zhang, Duojun Zhao, Jixiang Wang, and Gaoyuan
Sun for their excellent technical supports and contributions to this project.
We also thank Professor Jun Tan for his helpful comments and discussions.
Conceived and designed the experiments: XZ SH ISAM JY. Performed
the experiments: YF YY LP. Analyzed the data: YF HW TZ. Contributed
reagents/materials/analysis tools: MY XY. Wrote the paper: YF HW TZ.
1. Lang BF, Gray MW, Burger G (1999) Mitochondrial genome evolution and the
origin of eukaryotes. Annu Rev Genet 33: 351–397.
Li L, Wang B, Liu Y, Qiu YL (2009) The complete mitochondrial genome
sequence of the hornwort Megaceros aenigmaticus shows a mixed mode of
conservative yet dynamic evolution in early land plant mitochondrial genomes.
J Mol Evol 68: 665–678.
3. Wang B, Xue J, Li L, Liu Y, Qiu YL (2009) The complete mitochondrial genome
sequence of the liverwort Pleurozia purpurea reveals extremely conservative
mitochondrialgenomeevolutionin liverworts.Curr Genet 55: 601–609.
Brennicke A, Grohmann L, Hiesel R, Knoop V, Schuster W (1993) The
mitochondrial genome on its way to the nucleus: different stages of gene transfer
in higher plants. FEBS Lett 325: 140–145.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org11 May 2012 | Volume 7 | Issue 5 | e37164
5. Cummings MP, Nugent JM, Olmstead RG, Palmer JD (2003) Phylogenetic Download full-text
analysis reveals five independent transfers of the chloroplast gene rbcL to the
mitochondrial genome in angiosperms. Curr Genet 43: 131–138.
Wolfe KH, Li WH, Sharp PM (1987) Rates of nucleotide substitution vary
greatly among plant mitochondrial, chloroplast, and nuclear DNAs. Proc Natl
Acad Sci U S A 84: 9054–9058.
Kubo T, Newton KJ (2008) Angiosperm mitochondrial genomes and mutations.
Mitochondrion 8: 5–14.
McCauley DE, Olson MS (2008) Do recent findings in plant mitochondrial
molecular and population genetics have implications for the study of gynodioecy
and cytonuclear conflict? Evolution 62: 1013–1025.
Winkler M, Kuck U (1991) The group IIB intron from the green alga
Scenedesmus obliquus mitochondrion: molecular characterization of the in vitro
splicing products. Curr Genet 20: 495–502.
10. Hiesel R, Combettes B, Brennicke A (1994) Evidence for RNA editing in
mitochondria of all major groups of land plants except the Bryophyta. Proc Natl
Acad Sci U S A 91: 629–633.
11. Giege P, Brennicke A (1999) RNA editing in Arabidopsis mitochondria effects
441 C to U changes in ORFs. Proc Natl Acad Sci U S A 96: 15324–15329.
12. Palmer JD, Adams KL, Cho Y, Parkinson CL, Qiu YL, et al. (2000) Dynamic
evolution of plant mitochondrial genomes: mobile genes and introns and highly
variable mutation rates. Proc Natl Acad Sci U S A 97: 6960–6966.
13. Marienfeld J, Unseld M, Brennicke A (1999) The mitochondrial genome of
Arabidopsis is composed of both native and immigrant information. Trends
Plant Sci 4: 495–502.
14. Ward BL, Anderson RS, Bendich AJ (1981) The mitochondrial genome is large
and variable in a family of plants (cucurbitaceae). Cell 25: 793–803.
15. Sghaier-Hammami B, Valledor L, Drira N, Jorrin-Novo JV (2009) Proteomic
analysis of the development and germination of date palm (Phoenix dactylifera
L.) zygotic embryos. Proteomics 9: 2543–2554.
16. IS A-M (1996) Date palm. Arabian Global Encyclopedia 7: 182–187.
17. Al-Dous EK, George B, Al-Mahmoud ME, Al-Jaber MY, Wang H, et al. (2011)
De novo genome sequencing and comparative genomics of date palm (Phoenix
dactylifera). Nat Biotechnol 29: 521–527.
18. Yang M, Zhang X, Liu G, Yin Y, Chen K, et al. (2010) The complete
chloroplast genome sequence of date palm (Phoenix dactylifera L.). PLoS One 5:
19. Alverson AJ, Rice DW, Dickinson S, Barry K, Palmer JD (2011) Origins and
recombination of the bacterial-sized multichromosomal mitochondrial genome
of cucumber. Plant Cell 23: 2499–2513.
20. Wang D, Wu YW, Shih AC, Wu CS, Wang YN, et al. (2007) Transfer of
chloroplast genomic DNA to mitochondrial genome occurred at least 300 MYA.
Mol Biol Evol 24: 2040–2048.
21. Binder S, Marchfelder A, Brennicke A (1996) Regulation of gene expression in
plant mitochondria. Plant Mol Biol 32: 303–314.
22. Joyce CM, Steitz TA (1994) Function and structure relationships in DNA
polymerases. Annu Rev Biochem 63: 777–822.
23. Ong HC, Palmer JD (2006) Pervasive survival of expressed mitochondrial rps14
pseudogenes in grasses and their relatives for 80 million years following three
functional transfers to the nucleus. BMC Evol Biol 6: 55.
24. Kadowaki K, Kubo N, Ozawa K, Hirai A (1996) Targeting presequence
acquisition after mitochondrial gene transfer to the nucleus occurs by
duplication of existing targeting signals. EMBO J 15: 6652–6661.
25. Mower JP, Bonen L (2009) Ribosomal protein L10 is encoded in the
mitochondrial genome of many land plants and green algae. BMC Evol Biol
26. Kubo N, Arimura S (2010) Discovery of the rpl10 gene in diverse plant
mitochondrial genomes and its probable replacement by the nuclear gene for
chloroplast RPL10 in two lineages of angiosperms. DNA Res 17: 1–9.
27. Huh TY, Gray MW (1982) Conservation of ribosomal RNA gene arrangement
in the mitochondrial DNA of angiosperms. Plant Molecular Biology 1: 245–249.
28. Tian X, Zheng J, Hu S, Yu J (2007) The discriminatory transfer routes of tRNA
genes among organellar and nuclear genomes in flowering plants: a genome-
wide investigation of indica rice. J Mol Evol 64: 299–307.
29. Stern DB, Lonsdale DM (1982) Mitochondrial and chloroplast genomes of
maize have a 12-kilobase DNA sequence in common. Nature 299: 698–702.
30. Stern DB, Palmer JD (1984) Extensive and widespread homologies between
mitochondrial DNA and chloroplast DNA in plants. Proc Natl Acad Sci U S A
31. Alverson AJ, Wei X, Rice DW, Stern DB, Barry K, et al. (2010) Insights into the
evolution of mitochondrial genome size from complete sequences of Citrullus
lanatus and Cucurbita pepo (Cucurbitaceae). Mol Biol Evol 27: 1436–1448.
32. Covello PS, Gray MW (1989) RNA editing in plant mitochondria. Nature 341:
33. Gualberto JM, Lamattina L, Bonnard G, Weil JH, Grienenberger JM (1989)
RNA editing in wheat mitochondria results in the conservation of protein
sequences. Nature 341: 660–662.
34. Hiesel R, Wissinger B, Schuster W, Brennicke A (1989) RNA editing in plant
mitochondria. Science 246: 1632–1634.
35. Hoch B, Maier RM, Appel K, Igloi GL, Kossel H (1991) Editing of a chloroplast
mRNA by creation of an initiation codon. Nature 353: 178–180.
36. Wintz H, Hanson MR (1991) A termination codon is created by RNA editing in
the petunia atp9 transcript. Curr Genet 19: 61–64.
37. Shikanai T (2006) RNA editing in plant organelles: machinery, physiological
function and evolution. Cell Mol Life Sci 63: 698–708.
38. Wissinger B, Brennicke A, Schuster W (1992) Regenerating good sense: RNA
editing and trans splicing in plant mitochondria. Trends Genet 8: 322–328.
39. Pring D, Brennicke A, Schuster W (1993) RNA editing gives a new meaning to
the genetic information in mitochondria and chloroplasts. Plant Mol Biol 21:
40. Mower JP (2005) PREP-Mt: predictive RNA editor for plant mitochondrial
genes. BMC Bioinformatics 6: 96.
41. Howad W, Kempken F (1997) Cell type-specific loss of atp6 RNA editing in
cytoplasmic male sterile Sorghum bicolor. Proc Natl Acad Sci U S A 94:
42. Grosskopf D, Mulligan RM (1996) Developmental- and tissue-specificity of RNA
editing in mitochondria of suspension-cultured maize cells and seedlings. Curr
Genet 29: 556–563.
43. Tian X, Zheng J, Hu S, Yu J (2006) The Rice Mitochondrial Genomes and
Their Variations. Plant Physiology 140: 401–410.
44. Lilly JW, Havey MJ (2001) Small, repetitive DNAs contribute significantly to the
expanded mitochondrial genome of cucumber. Genetics 159: 317–328.
45. Alverson AJ, Zhuo S, Rice DW, Sloan DB, Palmer JD (2011) The mitochondrial
genome of the legume Vigna radiata and the analysis of recombination across
short mitochondrial repeats. PLoS One 6: e16404.
46. Clifton SW, Minx P, Fauron CM, Gibson M, Allen JO, et al. (2004) Sequence
and comparative analysis of the maize NB mitochondrial genome. Plant Physiol
47. Hedtke B, Legen J, Weihe A, Herrmann RG, Borner T (2002) Six active phage-
type RNA polymerase genes in Nicotiana tabacum. Plant J 30: 625–637.
48. Kuhn K, Weihe A, Borner T (2005) Multiple promoters are a common feature
of mitochondrial genes in Arabidopsis. Nucleic Acids Res 33: 337–346.
49. He S, Abad AR, Gelvin SB, Mackenzie SA (1996) A cytoplasmic male sterility-
associated mitochondrial protein causes pollen disruption in transgenic tobacco.
Proc Natl Acad Sci U S A 93: 11763–11768.
50. Fujii S, Toda T, Kikuchi S, Suzuki R, Yokoyama K, et al. (2011) Transcriptome
map of plant mitochondria reveals islands of unexpected transcribed regions.
BMC Genomics 12: 279.
51. Mortazavi A, Williams BA, McCue K, Schaeffer L, Wold B (2008) Mapping and
quantifying mammalian transcriptomes by RNA-Seq. Nat Methods 5: 621–628.
52. Carlsson J, Leino M, Sohlberg J, Sundstrom JF, Glimelius K (2008)
Mitochondrial regulation of flower development. Mitochondrion 8: 74–86.
53. Huang J, Struck F, Matzinger DF, Levings CS III (1994) Flower-enhanced
expression of a nuclear-encoded mitochondrial respiratory protein is associated
with changes in mitochondrion number. Plant Cell 6: 439–448.
54. Preuten T, Cincu E, Fuchs J, Zoschke R, Liere K, et al. (2010) Fewer genes than
organelles: extremely low and variable gene copy numbers in mitochondria of
somatic plant cells. Plant J 64: 948–959.
55. Kalantidis K, Wilson Z, Mulligan B (2002) Mitochondrial gene expression in
stamens is differentially regulated during male gametogenesis in Arabidopsis.
Sexual Plant Reproduction 14: 299–304.
56. Thomson MC, Macfarlane JL, Beagley CT, Wolstenholme DR (1994) RNA
editing of mat-r transcripts in maize and soybean increases similarity of the
encoded protein to fungal and bryophyte group II intron maturases: evidence
that mat-r encodes a functional protein. Nucleic Acids Res 22: 5745–5752.
57. Finnegan PM, Brown GG (1990) Transcriptional and Post-Transcriptional
Regulation of RNA Levels in Maize Mitochondria. Plant Cell 2: 71–83.
58. Homer N, Merriman B, Nelson SF (2009) BFAST: an alignment tool for large
scale genome resequencing. PLoS One 4: e7767.
59. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local
alignment search tool. J Mol Biol 215: 403–410.
60. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, et al. (1997) Gapped
BLAST and PSI-BLAST: a new generation of protein database search
programs. Nucleic Acids Res 25: 3389–3402.
61. Lowe TM, Eddy SR (1997) tRNAscan-SE: a program for improved detection of
transfer RNA genes in genomic sequence. Nucleic Acids Res 25: 955–964.
62. Gardner PP, Daub J, Tate JG, Nawrocki EP, Kolbe DL, et al. (2009) Rfam:
updates to the RNA families database. Nucleic Acids Res 37: D136–140.
63. Kurtz S, Choudhuri JV, Ohlebusch E, Schleiermacher C, Stoye J, et al. (2001)
REPuter: the manifold applications of repeat analysis on a genomic scale.
Nucleic Acids Res 29: 4633–4642.
64. Benson G (1999) Tandem repeats finder: a program to analyze DNA sequences.
Nucleic Acids Res 27: 573–580.
65. Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, et al. (2007)
Clustal W and Clustal X version 2.0. Bioinformatics 23: 2947–2948.
66. Castresana J (2000) Selection of conserved blocks from multiple alignments for
their use in phylogenetic analysis. Mol Biol Evol 17: 540–552.
67. Guindon S, Dufayard JF, Lefort V, Anisimova M, Hordijk W, et al. (2010) New
algorithms and methods to estimate maximum-likelihood phylogenies: assessing
the performance of PhyML 3.0. Syst Biol 59: 307–321.
68. Krzywinski M, Schein J, Birol I, Connors J, Gascoyne R, et al. (2009) Circos: an
information aesthetic for comparative genomics. Genome Res 19: 1639–1645.
69. Chaw SM, Shih AC, Wang D, Wu YW, Liu SM, et al. (2008) The
mitochondrial genome of the gymnosperm Cycas taitungensis contains a novel
family of short interspersed elements, Bpu sequences, and abundant RNA
editing sites. Mol Biol Evol 25: 603–615.
Date Palm Mitochondrial Genome and Transcriptome
PLoS ONE | www.plosone.org12 May 2012 | Volume 7 | Issue 5 | e37164