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SYMBIOGENETICS
New naturally transgenic plants: 2020 update
Tatiana Matveeva
Department of Genetics and Biotechnology, Saint Petersburg State University, Universitetskaya nab., 7-9, Saint Petersburg, 199034, Russian Federation
Address correspondence and requests for materials to Tatiana Matveeva, t.v.matveeva@spbu.ru
Abstract
Agrobacterium-mediated gene transfer leads to crown gall or hairy roots disease, due to expression of transferred T-DNA genes. Spontaneous plant regeneration from the transformed tissues can produce natural transformants carrying cellular T-DNA (cT-DNA) sequences of agrobacterial origin. In 2019, based on genomic sequencing data, cT-DNA horizontally transferred from Agrobacte-rium were found in two dozen species of angiosperms. This made it possible to evaluate the spread of this phenomenon, as well as make some generalizations regarding the diversity of horizontally transferred genes. The presented research is a continuation of work in this field. It resulted in the description of new naturally occurring transgenic species Aeschynomene evenia C.Wright, Eperua falcata Aubl., Eucalyptus cloeziana F. Muell., Boswellia sacra Flueck., Kewa caespitosa (Friedrich) Christenh., Pharnaceum exiguum Adamson, Silene noctíflora L., Nyssa sinensis Oliv., Vaccinium corymbosum L., Populus alba L. * Populus glandulosa Moench. The previously identified patterns regarding the frequency of the occurrence of natural transformants and the general properties of the cT-DNAs were confirmed in this study.
Keywords: cT-DNA, horizontal gene transfer, naturally-transgenic plants
Citation: Matveeva, T. 2021. New naturally transgenic plants: 2020 update. Bio. Comm. 66(1): 36-46. https://doi. org/10.21638/spbu03.2021.105
Authors' information: Tatiana Matveeva, Dr. of Sci. in Biology, Professor, orcid. org/0000-0001-8569-6665
Manuscript Editor: Kirill Antonets, Department of Cytology and Histology, Faculty of Biology, Saint Petersburg State University, Saint Petersburg, Russia
Received: September 24, 2020;
Revised: November 8, 2020;
Accepted: November 25, 2020.
Copyright: © 2021 Matveeva. This is an open-access article distributed under the terms of the License Agreement with Saint Petersburg State University, which permits to the authors unrestricted distribution, and self-archiving free of charge.
Funding: The article was made with support of the Ministry of Science and Higher Education of the Russian Federation in accordance with agreement № 075-15-2020922 dated 16.11.2020 on providing a grant in the form of subsidies from the federal budget of the Russian Federation. The grant was provided for state support for the creation and development of a world-class scientific center, Agrotechnologies for the Future.
Competing interests: The authors have declared that no competing interests exist.
Introduction
Agrobacterium-mediated transformation is the most common method for obtaining genetically modified plants. It is based on the ability of these soil bacteria to transfer a fragment of their plasmid (T-DNA, transferred DNA) and integrate it into the chromosome of the host plant. In nature, such a transfer leads to the development of two types of diseases: crown gall and hairy root diseases. These neoplasms are transgenic tissues on a non-transgenic plant. Scientists have managed to replace T-DNA genes with the sequences they need, transfer them using agrobacterial vectors into plant cells, and regenerate whole plants from such transgenic cells (Nester, 2014). It turned out that similar processes occur in nature, since plants were found to contain sequences homologous to the T-DNA of Agrobacterium in their genomes (Chen and Otten, 2017; Matveeva, 2018). This T-DNA was named cellular T-DNA (cT-DNA). The first such plants were found within the genus Nicotiana (White et al., 1983), and more than 20 years later in the genomes of Linaria and Ipomoea (Matveeva et al., 2012; Kyndt et al., 2015). Until 2019, the list of naturally transgenic plants was limited to these three genera. Digressing slightly from the main topic, we want to note that we are aware that the phylogeny of the genus Agrobacterium has been revised since the first discovery of T-DNA in wild plants (Young et al., 2001, 2003; Farrand et al., 2003); however, in the text of the manuscript we will use the collective term Agrobacterium as a tribute to tradition, and also because of the impossibility of accurately identifying the type of bacteria that participated in the transformation of the plant millions of years ago. The small fragments of T-DNA present in plant genomes are not sufficient for this. At the same time, further in the text of the manuscript, when indicating the closest of the modern strains, we will provide their modern name.
The development of genomic sequencing and bio-informatics methods have opened up new opportunities for the search for new natural GMOs. Such a search was crowned with success in 2019 (Matveeva and Otten, 2019): another two dozen species, the ancestors of which underwent Agrobacterium-mediated transformation during their evolution, were described within the genera Eu-trema, Arachis, Nissolia, Quillaja, Euphorbia, Parasponia, Trema, Humulus, Psidium, Eugenia, Juglans, Azadirachta, Silene, Dianthus, Vaccinium, Camellia and Cuscuta. Analysis of transcriptome data revealed an additional list of natural transformants. However, the degree of confidence in natural transgenicity based on transcriptomic data is lower than that based on results of genome sequencing and assembly. This is due to the lack of information about the localization site of the sequences, which leads to the possibility that the sequences result from Agrobacterium DNA contamination. The most interesting results of transcriptome assembly were several T-DNA-like sequences of the representatives of the genus Diospyros, containing a combination of opine and plast-genes. Matveeva and Otten's (2019) study was done exclusively using bioinfor-matic analysis of published sequences of plant genomes. A few months later, an article was published in which molecular methods confirmed the presence of T-DNA in plants of the genus Cuscuta, previously identified by bio-informatics means (Zhang et al., 2020). Numerous new examples of natural transformants show that at least 7 % of the dicotyledonous species are naturally transformed plants, and provide valuable material for studying the role of horizontal gene transfer in plant evolution (Matveeva and Otten, 2019). These results also serve as an important argument in support of GMOs.
A year has passed since the publication of Matveeva and Otten (2019). During this time, new plant genomes were sequenced and deposited in the NCBI database (O'Leary et al. 2016). The aim of this work was to update the list of naturally transgenic plants taking into account new NGS data, and generalize all the results obtained.
Material and methods
The search for T-DNA-like sequences was done based on National Center for Biotechnology Information (NCBI) Whole-Genome Shotgun (WGs) contigs of all plant genomes sequenced since April 2019 to date, using the TBLASTN algorithm with default settings. In the second step, Vir protein sequences were used to search for possible Agrobacterium contaminations in those genomes. In the third step, contigs that potentially encoded T-DNA-like protein sequences with identity levels 30 % or higher were analyzed further. They were used as queries in BLASTX with default settings to detect the closest protein homologs and to identify proteins encoded by plant genes surrounding the cT-DNA. All query
sequences are detailed in our previous paper (Matveeva and Otten, 2019). The Vector NTI AdvanceTM software was used to build the cT-DNA maps.
Phylogenetic analysis of rolB/C homologs was done in MEGA 7.0 (Kumar et al., 2016) by using the Maximum Likelihood method based on the JTT matrix-based model (Jones et al., 1992) (In addition, the Dayhoff matrix based model (Schwarz and Dayhoff, 1979), Poisson correction model (Zuckerkandl and Pauling, 1965) and Equal Input model (Tajima and Nei, 1984) were used for more reliable conclusions). The bootstrap consensus tree inferred from 500 replicates was taken to represent the evolutionary history of the taxa analyzed (Felsenstein, 1985). Branches corresponding to partitions reproduced in less than 50 % bootstrap replicates were collapsed. Initial tree(s) for the heuristic search were obtained automatically by applying Neighbor-Join and BioNJ algorithms to a matrix of pairwise distances estimated using a JTT model, and then selecting the topology with superior log likelihood value. The analysis involved 19 amino acid sequences. All positions with less than 95 % site coverage were eliminated. That is, fewer than 5 % alignment gaps, missing data, and ambiguous bases were allowed at any position. There were a total of 140 positions in the final dataset.
The supplementary materials present a similar analysis performed by UPGMA method (Sneath and Sokal, 1973) and neighbor-joining method (Saitou Nei, 1987).
Results and discussion
Since April 2019 (Matveeva and Otten, 2019), the genomes of another 206 angiosperm species have been sequenced. New examples of natural GMOs were identified in 10 species (about 5 %) from 10 genera, 9 families and 7 orders, according to the previously described methodology (Matveeva and Otten, 2019). They are listed in Table 1. Schemes of extended cT-DNAs are shown in Figure 1.
For representatives of two genera, the cT-DNA structure was specified. At the same time, their trans-genic nature was described earlier.
Until recently, two variants of cT-DNA have been characterized in plants of the genus Ipomoea (Kyndt et al., 2015; Quispe-Huamanquispe et al., 2019). In our study, based on the genome sequences of I. trifida (Kunth) G. Don and I. batatas (L.) Lam., a new cT-DNA variant was discovered. It contains mas2'-like and mas1 -like sequences. The fragment that we found in I. trifida was named It-TDNA3. A similar (86 %) fragment was also found in I. batatas. At the same time, the boundary sequences of plant origin are 97 % similar, showing that they result from the same transformation event. The database also contains short contigs containing mas2' homologues. However, it is not possible to attribute them to any extended sequence. Further research is required
Fig. 1. Structure of cT-DNA plant species. (Wide green arrows show sequences similar to Agrobacterium T-DNA genes, blue arrows show inverted repeats, green thin arrows show direct repeats. Red arrows show short repeating sequences).
to clarify the nature of these sequences. Therefore, they are not currently listed in the results table.
We predicted a cT-DNA in Diospyros lotus L. (date-plum) based on the analysis of the TSA database (Matveeva and Otten, 2019). Analysis of the results of genome assembly made it possible to describe seven variants of cT-DNA in this species, representing footprints of several independent transformation events in the evolution of
this species (Fig. 1). Dl-TDNA1 and 2 are located close to the boundaries of the assembled sequences. They share 99 % similarity and may be part of the same cT-DNA. If so, then this is the youngest cT-DNA in the genome of this species, which can be dated by the repeat structure. It is followed by Dl-TDNA5, 7 and 6. Dl-TDNA6 is the oldest one. Other traces of multiple acts of agrobacte-rial transformation in the evolution of ancestral forms of
S
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order > Species, cultivar, line, isolate Intact* Identity level to proteins from NCBI Similarity level between 2 arms of the cT-DNA
E ro M- Accession # Gene homolog position % of Identity Organism and protein ID
Populus alba x Populus glandulosa isolate Beijing Shi SMNX01000141.1 or/73-1 ike - 98346 -99995 49 BAB16132.1 Rhizobium rhizo-genes (Riker et al. 1930) Young etal. 2001 92
i/i CD TO CD ro/e -like - 102808-102180** 60 CAA82552.1 R. rhizogenes
JZ 00 'CL CD U TO U or/73-1 ike - 106748-104516 49 BAB16132.1 R. rhizogenes
ro 5 "ci U1 acs-I ike - 112142-112940 86 WPJ74054263.1 R. rhizogenes
Aeschynomene evenia isolate CIAT22838 RYYW01000009.1 mis-like + 25927086 — 25926118 62 WPJ74054202.1 R. rhizogenes n/a
Fabales TO CD U TO _Q TO Ll_ Eperua falcata CWNJ01257379.1 nos-like + 1339-149 54 WP_167693616.1 Sinorhizo-bium meliloti (Dangeard 1926) De Lajudie etal. 1994 n/a
Eucalyptus cloeziana isolate ANBG68772 JABKB0010000005.1 mas 7'-like - 19003305 — 19004414 70 P27874.2 R. rhizogenes 95
ags-llke - 19005239-19004732 69 WP_172690594.1 Agrobacte-rium sp.
mas2-llke - 19029872-19028766 61 AIM40180.1 R. rhizogenes
mos7'-like - 19030390-19031235 19046200-19046439 71...47 P27874.2 R. rhizogenes
ags-llke - 19047360-19046844 77 WPJB2488587.1 Agrobacte-rium sp.
mas2-llke - 19064603 — 19063528 61 AIM40180.1 R. rhizogenes
mas7'-like - 19065068 — 19066309 66 WP_034520976.1 Agrobacte-rium sp.
i/i CD CD TO CD U mas2-llke - 19091525 — 19090591 44 WP_172690593.1 Agrobacte-rium sp.
-CT5 >1 5 -CT5 >1 5 mas2-llke - 19097721 -19096760 56 (WP_172690593.1 Agrobacte-rium sp.
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Continuation of the Table 1
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order Species, cultivar, line, isolate Intact* Identity level to proteins from NCBI Similarity level between 2 arms of the cT-DNA
£ £ Accession # Gene homolog position % of identity Organism and protein ID
Boswellia sacra isolate BS-S2 SNVD01001790.1 rolD - 4554 — 5602 69 WP_034521016.1 Agrobacte-rium sp. n/a
iaaM-Wke - 15557-13686 47 WP_174054196.1 R.rhizogenes 87
iaaM-Wke - 17274— 19107 45 WP_174054196.1 R.rhizogenes
or/70-llke - 19840 -20193 40 AAA22094.1 R.rhizogenes n/a
CD ro CL) U ro HAD hydrolase family 7 21778 — 22257 40 WP_149743959.1 Rhizobium sp.
ro TB-orfU-2- like - 23456 — 23151 31 AIM40179.1 R.rhizogenes
c 'CL fD CD cu to D CO TB-orfU- 7-like - 23561 -23151 29 AIM40178.1 Nicotiana tomen-tosiformis
Kewa caespitosa ONZAOI 007622.1 m/'s-llke + 509 — 1510 65 BAB85949.1 Nicotiana glauca n/a
Kewaceae ONZAOI013126.1 m/'s-llke - 1503 — 2552 62 WP_176453671.1 R.rhizogenes n/a
ONZAOI 022047.1 m/'s-llke - 1924 — 2417 54 WP_176453671.1 R.rhizogenes n/a
ONZAOI 003306.1 m/'s-llke - 4964 — 5356 51 BAB85949.1 Nicotiana glauca n/a
c '&0 ^ CL) Pharnaceum exiguum ONZKOI002102.1 m/'s-llke + 26894 — 27826 66 BAB85949.1 Nicotiana glauca n/a
ONZKOI 003836.1 m/'s-llke + 7318 — 8232 62 BAB85949.1 Nicotiana glauca 73
% s .> u m/'s-llke - 14903 — 15822 59 BAB85949.1 Nicotiana glauca
Silene noctíflora Isolate OPL-1.1 VHZZ01000004.1 cus-1 Ike - 1170610 — 1176104 60 WP_156551602.1 Allorhizobium vitis (Ophel and Kerr 1990) n/a
cus-1 Ike + 1182125 — 1183057 64 WP_156551602.1 A. vitis n/a
cus-1 Ike + 1210326 — 1209418 64 WP_156551602.1 A. vitis n/a
VHZZ01016788.1 cus-1 Ike + 37722 — 36790 63 WP_174084799.1 A. vitis n/a
VHZZ01000003.1 cus-llke - 5037 — 5411 54 WP_174084799.1 A. vitis n/a
cu TO cus-llke - 25613-24951 68 WP_174084799.1 A. vitis n/a
U cus-llke + 28697 -27765 63 WP_174084799.1 A. vitis n/a
CL O J>> cus-llke + 35431 -34499 62 WP_174084799.1 A. vitis n/a
IV U VHZZ01056725.1 cus-llke - 6031 —5510 72 WP_071208191.1 A. vitis n/a
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order > Species, cultivar, line, isolate Intact* Identity level to proteins from NCBI Similarity level between 2 arms of the cT-DNA
E >2 Accession # Gene homolog position % of Identity Organism and protein ID
Cornales Nys- sa- ceae Nyssa sinensis isolate J 267 VIRR01000271.1 rolB/C- like + 5364628 -5365320 52 XP_001881215.1 Laccaria bicolor n/a
i/i cu Diospyros lotus cv. Kun-senshl D/-T-DNA1 BEWH01006414.1 cus-1 Ike - 41404 — 42306 73 WP_156551602.1 A. vitis n/a
u LU or/74-llke - 44357 -43779 56 WP_174054201.1 R. rhizogenes
DI-T-DNA2 BEWH01000237.1 cus-1 Ike + 334877 — 335779 73 WP_156551602.1 A. vitis n/a
or/74-llke - 337830 -337252 56 WP_174054201.1 R. rhizogenes
acs-1 Ike + 347771 — 348979 81 GAJ95539.1 R. rhizogenes
DI-T-DNA3 BEWH01006419.1 acs-1 Ike + 8509 — 7220 80 GAJ95539.1 R. rhizogenes n/a
orf3-like - 10262 — 11610 70 KEA04445.1 R. rhizogenes
IS5 transposase - 12185-12934 83 WP_184141638.1 Shinella fusca Vaz-Morelra et al. 2010
iaaM-Wke - 13558-15830 45 WP_034521028.1 Agrobacte-rium sp.
DI-T-DNA4 BEWH01000029.1 orf8-like + 2523369 — 2522746 68 WP_116979321.1 Agrobacterium salinitolerans Yan et al. 2017 n/a
or/3-llke - 2525305 -2523957 72 KEA04445.1 R. rhizogenes
orf2-like - 2525577 -2526392 WP_174054193.1 R. rhizogenes
acs-1 Ike - 2526938 — 2528244 73 KEA04447.1 Agrobacterium sp.
DI-T-DNA5 BEWH01004217.1 sus-llke + 21622 — 22629 77 WP_174080856.1 R. rhizogenes 97
IS630 - 25887 -26667 83 WP_165826447.1 Rhizobium wuzhouense Yuan et al. 2018
C-like - 28060 -26786 76 WP_174054195.1 R. rhizogenes
C-like - 30052 — 31252 76 WP_174054195.1 R. rhizogenes
IS630 - 32218-31443 83 WP_165826447.1 R. wuzhouense
sus-llke - 36499 -35486 76 WP_174080856.1 R. rhizogenes
End of the Table 1
■t*
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order family Species, cultivar, line, isolate Accession # Gene homolog Intact* position Identity level to proteins from NCBI Similarity level between 2 arms of the cT-DNA
% of identity Organism and protein ID
Ebenaceae DI-T-DNA5a BEWH01000041.1 (2128496 — 214 3594) 99% Identical to BEWH01004217.1
DI-T-DNA6 BEWH01000056.1 sus-llke - 1177021 -1177736 58 WP_034521016.1 Agrobacte-rium sp 91
or/57 7-llke - 1179590-1178145 60 AIM40183.1 Nicotiana tomen-tosiformis
or/74-llke - 1180901 — 1180354 68 AIM40184.1 Nicotiana tomen-tosiformis
IS3 family trans-posase - 1182902 — 1181706 70 WP_143239454.1 Agrobacte-rium rosae
sus-llke - 1183310 — 1184002... 1185015-1184824 47 WP_174080856.1 R. rhizogenes
IS3 family trans-posase - 1185301 -1186508 73 WP_143239454.1 Agrobacte-rium rosae
or/57 7-llke - 1187393-1187790 60 AIM40183.1 Nicotiana tomen-tosiformis
sus-llke - 1189225-1188221 55 WP_174080856.1 R. rhizogenes
ocs-llke - 1190796 — 1189840 38 KIJ92238.1 Laccaria amethys-tina
DI-T-DNA7 BEWH01000037.1 or/73-llke - 262687 -264989 50 WP_174075801.1 R. rhizogenes 92
or/73-llke - 271357 -270518... 269030 — 267893 48 WP_174075801.1 R. rhizogenes
Vac- cinioi- deae Vaccinium corymbosum cultivar W8520 JACAOBOI 0009726.1 rolB/C- like + 1 —729 41 XP_001884861.1 Laccaria bicolor n/a
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order family Specles, cultivar, Une, isolate Accession # Gene homolog Intact* position Identity level to proteins from NCBI Similarity level between 2 arms of the cT-DNA
% of Identity Organism and protein ID
(is this similar to the earlier Silene T-DNA?)Solanales Convolvuláceae Ipomoea trífida cultivar Y22 SMMV01000602.1 Contains /¿-TDNA2
/C-TDNA3 SMMV01000003.1 mas2'- like - 13842355-13841152 71 WP_032488585.1 Agrobacte-rium sp. n/a
mas2-llke' - 13842876-13842715 72 AIM40180.1 R.rhizogenes
mosî'-llke - 13845955-13846602 77 P27874.2 R. rhizogenes
mosî'-llke - 13847831 — 13848160 75 P27874.2 R. rhizogenes
Ipomoea batatas cultivar Talzhong6 NXFB01008336.1 FLTB01041015.1 Contains Ab-TDNA1, described by Kyndt et al. (2015)
NXFB01000007.1 Contains /¿-TDNA2, described by Kyndt et al. (2015)
/¿-TDNA3 NXFB01000244.1 mas 7'-like - 54319-53674 77 WP_034520976.1 Agrobacte-rium s p. n/a
mas2'-llke - 56951 —58017 68 WP_032488585.1 Agrobacte-rium s p.
NXFB01000002.1 mas2'-llke - 27628510 — 27627275 63 AIM40180.1 R.rhizogenes n/a
* does not contain premature stop codons and / or frame shift Mr gene location on the negative strand
UJ
90
91
98
95
100
Laccaria bicolor rolBjC-like XP 001884963.1 Laccaria bicolor rolBjC-like XP_001884962.1 Laccaria bicolor rolBIC-like XP_001884964.1 Laccaria bicolor rolB IC-like XP_001881215.1 Laccaria bicolor rolB IC-like XP_001884861.1 -Nyssa sinensis rolBjC-like -Vaccinium macrocarpon rolBjC-like -Ensifersp. YR511ro/B/C-like SDN7877.1
100
97
90
-Linaria vulgaris rolC
-Nicotiana glauca rolC
-Rhizobium rhizogenes pRi8196 rolC
-Rhizobium rhizogenes pRil724 rolC
-Rhizobium rhizogenes pRi2659 rolC
-Rhizobium rhizogenes pRiA4 rolC
-Ipomoea batatas rolB/ C-like
-Rhizobium rhizogenes pRiA4 rolB
-Rhizobium rhizogenes pRil724 rolB
-Rhizobium rhizogenes pRi2659 rolB
-Rhizobium rhizogenes pRiA4 orfl3
Fig. 2. Molecular phylogenetic analysis of rolB/C homologs from Rhizobium, Ensifer, Laccaria, Ipomoea, Vaccinium and Nyssa species by Maximum Likelihood method based on the JTT matrix-based model. (Dayhoff matrix based model, Poisson correction model and Equal Input model resulted to the same topology of the tree). The cluster containing new rolB/C-like gene is outlined in red.
100
97
94
modern species have been previously described within the genera Nicotiana and Parasponia (Chen et al., 2014; Matveeva and Otten, 2019)
All new species of naturally transgenic plants belong to the same orders where natural GMOs were previously described. Vaccinium corymbosum L. and Silene noctiflora L. belong to genera in which natural GMOs were previously found. They contain sequences similar to those described earlier, which can be further used for phylogenetic studies based on the T-DNA structure. Our study also confirms the prevalence of opine genes in natural transformants. As before, we observe extended cT-DNAs organized as repeats. Inverted repeats may be generated during the process of T-DNA transfer and integration into plant chromosomes. Direct repeats may possibly be explained by DNA rearrangements associated with transposons found around the repeated cT-DNA regions. An interesting feature of eucalyptus T-DNA is that relatively short fragments of agrobacte-rial origin with similar opine genes are interspersed with extended DNA fragments of plant origin. A large number of repeats of the same opine genes, that are found in Silene species, Kewa caespitosa (Friedrich) Christenh. and Pharnaceum exiguum Adamson is another feature that requires further study; it may result from the insertion of multiple copies during the initial transformation
event, or from amplification of integrated copies at a later stage.
The data on the fine structure of cT-DNA in representatives of different taxa obtained earlier and in the present work can be further used to search for patterns of host specificity of modern agrobacterial strains. This issue can be investigated both from a phylogenetic and from an ecological point of view, since the idea of coevo-lution of symbionts is gaining in importance (Matveeva et al., 2018). We can already illustrate this thesis with the case of an unusual plast gene, which we described for the first time in the genomic sequence of Vaccinium macrocarpon Aiton. This fragment attracted our interest because it was closer to fungal plast-genes than agrobac-terial ones. In the present work, a similar sequence was found in Nyssa sinensis Oliv. Figure 2 shows that Nyssa, Vaccinium and Laccaria sequences cluster together with rolB/C-like gene of Ensifer sp. from the Rhizobiaceae family. Phylogenetic trees constructed by other methods (Supp. Fig. 1) have a similar topology, which confirms the reliability of this cluster. The genera Nyssa and Vac-cinium are not related, but these plants share similar habitats, characterized by increased moisture (https:// www.hortweek.com; Song and Hancock, 2011). Perhaps the search for an Agrobacterium strain similar to those that transformed these species will lead to the discovery
of bacterial determinants that are important for the survival of such strains in wet habitats.
Conclusion
Thus, in this study, new natural GMOs were described in 10 species (Aeschynomene evenia, Eperua falcate, Eucalyptus cloeziana, Boswellia sacra, Kewa caespitosa, Phar-naceum exiguum, Silene noctiflora, Nyssa sinensis, Vac-cinium corymbosum, Populus alba x Populus glandulosa) belonging to 10 genera, 9 families and 7 orders. The new type of cT-DNA was described in Ipomoea trifida and Ipomoea batatas, and the structure of cT-DNAs of Dio-spyros lotus cv. Kunsenshi was clarified. The previously identified patterns regarding the frequency of the occurrence of naturally transgenic plants and the general properties of the cT-DNAs were confirmed. The data obtained can be used further for genetic engineering, plant phylogeny and evolutionary research.
Acknowledgments
The author expresses her deep gratitude to Prof. L.Otten (IBMP, France) for critical reading of the manuscript, advice and comments.
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SUPPLEMENTS
Supplementary
Comparison of the topology of phylogenetic trees of rolB/C homologs constructed by A — Maximum Likelihood method based on the JTT matrix-based model (as in fig. 1) B — Neighbor-joining method based on the JTT matrix-based model С — UPGMA method based on the Poisson correction model D — UPGMA method based on the JTT matrix-based model
