Extensive length variation in the cpDNA trnT-trnF region of hemiparasitic Pedicularis and its phylogenetic implications

The cpDNA trnT-trnF region, a molecular marker widely used in the phylogenetic reconstruction at lower taxonomic levels, is relatively conserved in size and structure. In this region single length variation over 100 bp is much less common than small deletion for congeneric species of angiosperms. Here we examined evolutionary patterns of the trnT-trnF region in 43 species of Pedicularis, a species-rich genus with adaptive radiation. Four independent large deletions, varying from 203 to 297 bp in length, were detected from nine species of the genus, which might result from slipped-strand mispairing. These deletions occurred in different locations of the cpDNA region and in different clades of the phylogenetic tree, indicating that the deletion of large cpDNA fragments may be very frequent in the hemiparasitic lineage of the family Orobanchaceae. Parsimony analyses showed that section Cyathophora of Pedicularis, endemic to the Sino-Himalayan region, was a strongly supported monophyletic group. This section could have a recent origin followed by rapid radiation, considering that it is characterized by a large deletion in the trnT-trnF region and a relatively low interspecific sequence divergence.     

Phylogeny of Androcymbium (Colchicaceae) based on morphology and DNA sequences

The inclusion of species of Colchicum within Androcymbium in a previous cpDNA phylogeny of the Colchicaceae, questioned the monophyly of Androcymbium, and it was proposed to unite the two genera in Colchicum. Here we expand the previous phylogenetic analyses of Androcymbium by increasing the taxon sampling and adding more data. The analysis include 29 of the 57 species of Androcymbium, more cpDNA (trnL intron, trnL-trnF IGS, trnY-trnD IGS, and trnH-psbA IGS), and nDNA (RNApol2 intron 23) regions, and morphological and life-history traits data. Both parsimony and Bayesian inference were used. According to our data there is no reason to expand Colchicum to include Androcymbium, but we support the inclusion of Bulbocodium and Merendera within Colchicum. Morphology and life history traits are the main arguments in favor of recognizing Androcymbium as a well-circumscribed genus. In the phylogeny two Androcymbium groups are clearly differentiated: (1) one including species from Western and Eastern South Africa, Namibia, and North Africa, and (2) one including species from the north west of South Africa and south of Namibia.     

Species-diagnostic markers in <Emphasis Type="Italic">Larix</Emphasis> spp. based on RAPDs and nuclear,cpDNA, and mtDNA gene sequences,and their phylogenetic implications

Genetic markers from the nuclear, chloroplast, and mitochondrial genomes were developed to distinguish unambiguously among four larch species [Larix laricina (Du Roi) K. Koch, Larix decidua (Mill.), Larix kaempferi (Lamb.) Sarg., and Larix sibirica (Ledeb.)] used in intensive forestry in eastern North America. Nine random amplified polymorphic DNA (RAPD) fragments had good diagnostic value, and 3 out of 12 nuclear genes were found to harbor fixed interspecific polymorphisms implicating a total of 17 single nucleotide polymorphisms (SNPs) and 2 indels. The sequencing of five mtDNA introns (cox1-intron1, matR-intron1, nad1-intron b/c, nad3-intron1, and nad5-intron1) and four cpDNA regions (matK, trnL-intron, trnT–trnL and trnL–trnF intergenic spacers) resulted in the identification of 14 sites with fixed interspecific differences among the four species. Including the ten Larix species, one polymorphic site per 47 nucleotide sites sampled was observed for nuclear genes, one per 283 sites for cpDNA, and one per 374 sites for mtDNA. The phylogeny of the genus could be estimated from variation among the ten species detected in two cpDNA intergenic regions and four mtDNA introns. There was congruence between cpDNA and mtDNA phylogenies with three large groups delineated: the North American, North Eurasian, and South Asian taxa. The position of L. sibirica differed between organelle genomes. It was regrouped with South Asian species on the cpDNA tree, but with its North Eurasian congenerics on the mtDNA tree. To simplify the detection of diagnostic DNA sequence polymorphisms among the four main Larix species, cleaved amplified polymorphic sequence (CAPS) assays were developed from the polymorphisms identified in the various genomes. Seventeen primer–enzyme combinations were tested, and six were selected for their high level of informativeness. These new species-specific diagnostic markers should be useful for the certification of larch breeding materials and hybrid stocks used in intensive forestry in the northern hemisphere.     

Phylogenetics and taxonomic delimitation of the genus Guizotia (Asteraceae) based on sequences derived from various chloroplast DNA regions

Parsimony-based phylogenetic analyses of the genus Guizotia were undertaken based on DNA sequence data from the following chloroplast DNA (cpDNA) regions: trnT-trnL, trnL-trnF, trnY-rpoB, trnC-petN, psbM-trnD and rps16-trnQ intergenic spacers, trnL, rps16 and matK-5′trnK introns and matK gene. Out of the 26 primers used in this study, 14 were newly designed. The study was conducted to determine (1) the closest relative of Guizotia abyssinica, (2) the taxonomic status of some Guizotia taxa and (3) the subtribal placement of Guizotia in the tribe Heliantheae. The analyses of the sequence data showed that G. abyssinica, G. scabra ssp. scabra, G. scabra ssp. schimperi and G. villosa are phylogenetically closely related. However, G. scabra ssp. schimperi appeared as the most closely related taxon to G. abyssinica. Based on this phylogenetic analysis, we suggest that the two subspecies of G. scabra are better treated as separate species. The analysis also clearly demonstrated that “Chelelu” and “Ketcha” are distinct Guizotia species. The trnT-trnL and trnL-trnF intergenic spacer-based phylogenetic analysis of various subtribes of the tribe Heliantheae strongly supports the placement of the genus Guizotia within the subtribe Milleriinae.     

Phylogeny of Castanea (Fagaceae) based on chloroplast trnT-L-F sequence data

Species in the genus Castanea are widely distributed in the deciduous forests of the Northern Hemisphere from Asia to Europe and North America. They show floristic similarity but differences in chestnut blight resistance especially among eastern Asian and eastern North American species. Phylogenetic analyses were conducted in this study using sequences of three chloroplast noncoding trnT-L-F regions. The trnT-L region was found to be the most variable and informative region. The highest proportion of parsimony informative sites, more and larger indels, and higher pairwise distances between taxa were obtained at trnT-L than at the other two regions. The high A+T values (74.5%) in the Castanea trnT-L region may explain the high proportion of transversions found in this region where as comparatively lower A+T values were found in the trnL intron (68.35%) and trnL-F spacer (70.07%) with relatively balanced numbers of transitions and transversions. The genus Castanea is supported as a monophyletic clade, while the section Eucastanon is paraphyletic. C. crenata is the most basal clade and sister to the remainder of the genus. The three Chinese species of Castanea are supported as a single monophyletic clade, whose sister group contains the North American and European species. There is consistent but weak support for a sister–group relationship between the North American species and European species.     

Internal transcribed spacer region evolution in Larix and Pseudotsuga (Pinaceae)

The nuclear ribosomal DNA (nrDNA) internal transcribed spacer (ITS) region has been characterized in the sister genera Larix and Pseudotsuga (Pinaceae). Complete sequences were obtained for seven species of Larix from North America and Eurasia and five species of Pseudotsuga from western North America and eastern Asia. ITS region lengths ranged from 1759 to 1770 bp in Larix and from 1564 to 1571 bp in Pseudotsuga. In both genera, ITS1 is three times as long as the 5.8S plus ITS2 and contains subrepeats as observed in other genera of Pinaceae. Secondary structure models predicted that the subrepeats fold into terminal stem and loop domains. ITS polymorphism detected within individuals of Larix and Pseudotsuga suggests a slow rate of concerted evolution among nrDNA loci. Except for the placement of L. sibirica, phylogenetic analyses of the ITS region agreed with previously reported restriction site analyses of Larix and Pseudotsuga. The data were not consistent with phylogenetic hypotheses for Larix based primarily upon ovulate cone characters, failing to support a derivation of the North American L. laricina from a short-bracted Eurasian lineage. The phylogenetic hypothesis did not conflict with a stepping stone model of evolution for Pseudotsuga, but a basal lineage could not be inferred for either genus.     

Evolution of trnF(GAA) pseudogenes in cruciferous plants

In several studies we used the 5′-trnL(UAA)–trnF(GAA) region of the chloroplast DNA for phylogeographic reconstructions, gene diversity calculations and phylogenetic analyses among the genera Arabidopsis and Boechera. Despite the fact that extensive gene duplications are rare within the chloroplast genome of higher plants, within several genera of the Brassicaceae the anticodon domain of the trnF(GAA) gene exhibit extensive gene duplications with 1–12 tandemly repeated copies in close 5′-proximity of the functional gene. A recent re-examination and additional analysis of trnL(UAA)–trnF(GAA) regions from numerous cruciferous taxa not only reveal extensive trnF gene duplications, but also favour the hypothesis that in cruciferous taxa at least four independent phylogenetic lineages are characterized by these pseudogenes. Among these lineages there is one major clade of taxa carrying pseudogenes indicating an ancient split in crucifer evolution. In two case studies, Boechera and Arabidopsis, intra- and inter-molecular recombinations have been shown to be the reason for the reciprocal exchange of several similar motifs. However, functional constraints might favour two to three or five to six copies as shown for Arabidopsis and Boechera. Herein, we compare the occurrence and distribution of pseudogene copy number in the framework of a comprehensive survey of cpDNA haplotype variation in Boechera, the former genus Cardaminopsis and Arabidopsis thaliana and comment on the value of such kind of mutations in phylogenetic and evolutionary reconstructions.     

Molecular cytogenetic analysis of Siberian Larix species by fluorescence in situ hybridization

Karyotypes of three Larix species (L. sibirica, L. gmelinii, and L. cajanderi) were analyzed using fluorescence in situ hybridization (FISH) with 45S and 5S ribosomal RNA gene probes and 4′,6-diamidino-2-phenylindole (DAPI) staining. Two major 45S ribosomal DNA (rDNA) loci (per haploid genome) have been observed in the intercalary regions of two metacentric chromosomes, III and IV, of L. sibirica; in addition to them, minor nucleolus organizing regions (NORs) were mapped in pericentromeric regions of chromosomes I, II, VI, and XII. Two closely related species, L. gmelinii and L. cajanderi, showed similar hybridization patterns; both species possessed an additional major locus of 45S rDNA in the distal region of the long arm of submetacentric chromosome VII that is absent in L. sibirica. Only one locus of the 5S rDNA was found in all larch species we studied; it was located in the distal region of the chromosome III short arm, which also carried the major NOR in the opposite arm. This chromosome containing major loci of the two ribosomal RNA gene families can serve as a marker of the genus Larix. The intra- and interspecific karyotype diversity in the genus Larix is discussed.     

Molecular systematics, character evolution, and pollen morphology of Cistus and Halimium (Cistaceae)

Pollen analysis and parsimony-based phylogenetic analyses of the genera Cistus and Halimium, two Mediterranean shrubs typical of Mediterranean vegetation, were undertaken, on the basis of cpDNA sequence data from the trnL-trnF, and trnS-trnG regions, to evaluate limits between the genera. Neither of the two genera examined formed a monophyletic group. Several monophyletic clades were recognized for the ingroup. (1) The ??white and whitish pink Cistus??, where most of the Cistus sections were present, with very diverse pollen ornamentations ranging from striato-reticulate to largely reticulate, sometimes with supratectal elements; (2) The ??purple pink Cistus?? clade grouping all the species with purple pink flowers belonging to the Macrostylia and Cistus sections, with rugulate or microreticulate pollen. Within this clade, the pink-flowered endemic Canarian species formed a monophyletic group, but with weak support. (3) Three Halimium clades were recovered, each with 100% bootstrap support; all Halimium species had striato-reticulate pollen. Two Halimium clades were characterized by yellow flowers, and the other by white flowers.     

Phylogenetic relationships and intraspecific diversity of a North Patagonian Fescue: evidence of differentiation and interspecific introgression at peripheral populations

Peripheral sites usually offer suboptimal conditions for species with wide distributions, where expression of phenotypic variability and potential interspecific hybridization might be enhanced. The Patagonian steppe, the largest and southernmost dryland ecosystem in South America, is characterized by natural rangelands dominated by grasses. Festuca pallescens is a keystone species with a wide distribution in Patagonia over diverse environments reaching the extreme arid zones in the Somuncura plateau, a biogeographical island. Our aim is to study the phylogenetic relationships among Festuca pallescens populations as well as between this species and the sympatric F. argentina in North Patagonia. We analysed fourteen populations along a west-east transect of about 500 km in North Patagonia with three types of molecular markers: ITS, chloroplast trnL-F and eight nuclear microsatellites. Bayesian inferences, maximum parsimony and maximum likelihood analyses with trnL-F and ITS showed that F. pallescens is related to the Patagonian clade within the Festuca phylogeny. However, the easternmost populations of F. pallescens at Somuncura plateau were highly differentiated from the other populations and clustered with F. argentina (a sympatric species of the Asian-American clade). Principal coordinates analyses and Bayesian clustering performed with nuclear microsatellites as well as morphoanatomical traits, showed an intermediate position of one of these easternmost populations with respect to the two species, suggesting admixture. The high genetic variability observed in these peripheral populations highlight their relevance for conservation and might be indicating the existence of evolutionary processes triggering events of speciation in the Patagonian fescues.     

The complex history of the genus Ilex L. (Aquifoliaceae): evidence from the comparison of plastid and nuclear DNA sequences and from fossil data

 A plastid phylogeny of the genus Ilex based on three different loci (the atpB-rbcL spacer, trnL-trnF and rbcL) is compared with its nuclear phylogeny based on two different loci (the ribosomal ITS and the 5S RNA spacer). These two sets of molecular data are then compared to geographical and temporal data from the fossil record. The plastid phylogeny is strongly correlated with the geographic distribution of extant species. However, the nuclear phylogeny is strongly incongruent with the plastid phylogeny, suggesting frequent interlineage hybridizations. Moreover, the comparison of the ribosomal ITS tree and the 5S RNA spacer tree indicates also possible lineage sorting. Particularly interesting is the finding of two different Ilex lineages in the plastid American clade showing different biogeographic patterns in South America. One of them has a simple North American/South American biogeographical relationship. The other has complex biogeographical relationships, some species showing direct Asian/South American biogeographical relationships. During its history, the genus Ilex probably experienced frequent lineage sorting and interlineage hybridization with subsequent nuclear or cytoplasmic introgression, making the study of its history very complex. Received September 24, 2001; accepted August 19, 2002 Published online: November 28, 2002 Addresses of the authors: Jean-Fran?ois Manen (e-mail: manen@cjb.ville-ge.ch), Yamama Naciri-Graven, Conservatoire et Jardin Botaniques, Impératrice 1, CH-1292 Chambésy/Genève, Switzerland. Michael C. Boulter, Palaeobiology Research Unit, University of East London, Romford Road, London E15 4LZ, UK.     

Phylogeny of Carpinus and subfamily Coryloideae (Betulaceae) based on chloroplast and nuclear ribosomal sequence data

Phylogenetic studies were conducted for Carpinus and the subfamily Coryloideae (Betulaceae) using sequences of the chloroplast matK gene, the trnL-trnF region (trnL intron, and trnL [UAA] 3' exon-trnF [GAA] intergenic spacer) and the psbA-trnH intergenic spacer, and the nuclear ribosomal ITS regions. The combined analyses of the three chloroplast regions suggest that Coryloideae is monophyletic; Ostryopsis is sister to the Carpinus - Ostrya clade; Corylus is monophyletic and sister to the Ostrya - Carpinus - Ostryopsis clade; Ostrya is paraphyletic; and within Carpinus, species of sect. Carpinus from eastern Asia form a monophyletic group, whereas the positions of C. betulus from Europe and C. caroliniana from eastern North America are unresolved within the Carpinus clade. The cpDNA tree generated in this study is largely congruent with the previously published ITS results, but the ITS tree places Carpinus sect. Distegocarpus as sister to the Ostrya - Carpinus sect. Carpinus clade. Future work is needed to examine the relationships within the Ostrya - Carpinus clade, evaluate the generic status of Ostrya, and test the phylogenetic position of Ostryopsis.     

Molecular phylogeny of the subgenus Ceratotropis (genus Vigna, Leguminosae) reveals three eco-geographical groups and Late Pliocene-Pleistocene diversification: evidence from four plastid DNA region sequences

Background and Aims

The subgenus Ceratotropis in the genus Vigna is widely distributed from the Himalayan highlands to South, Southeast and East Asia. However, the interspecific and geographical relationships of its members are poorly understood. This study investigates the phylogeny and biogeography of the subgenus Ceratotropis using chloroplast DNA sequence data.

Methods

Sequence data from four intergenic spacer regions (petA-psbJ, psbD-trnT, trnT-trnE and trnT-trnL) of chloroplast DNA, alone and in combination, were analysed using Bayesian and parsimony methods. Divergence times for major clades were estimated with penalized likelihood. Character evolution was examined by means of parsimony optimization and MacClade.

Key Results

Parsimony and Bayesian phylogenetic analyses on the combined data demonstrated well-resolved species relationships in which 18 Vigna species were divided into two major geographical clades: the East Asia–Southeast Asian clade and the Indian subcontinent clade. Within these two clades, three well-supported eco-geographical groups, temperate and subtropical (the East Asia–Southeast Asian clade) and tropical (the Indian subcontinent clade), are recognized. The temperate group consists of V. minima, V. nepalensis and V. angularis. The subtropical group comprises the V. nakashimae–V. riukiuensis–V. minima subgroup and the V. hirtella–V. exilis–V. umbellata subgroup. The tropical group contains two subgroups: the V. trinervia–V. reflexo-pilosa–V. trilobata subgroup and the V. mungo–V. grandiflora subgroup. An evolutionary rate analysis estimated the divergence time between the East Asia–Southeast Asia clade and the Indian subcontinent clade as 3·62 ± 0·3 million years, and that between the temperate and subtropical groups as 2·0 ± 0·2 million years.

Conclusions

The findings provide an improved understanding of the interspecific relationships, and ecological and geographical phylogenetic structure of the subgenus Ceratotropis. The quaternary diversification of the subgenus Ceratotropis implicates its geographical dispersal in the south-eastern part of Asia involving adaptation to climatic condition after the collision of the Indian subcontinent with the Asian plate. The phylogenetic results indicate that the epigeal germination is plesiomorphic, and the germination type evolved independently multiple times in this subgenus, implying its limited taxonomic utility.     

Phylogeny of the Madagascan endemic family Didiereaceae

A molecular phylogeny of the Didiereaceae was produced through parsimony analysis of chloroplastrpl16 intron andtrnL-trnF andtrnT-trnL intergenic spacer sequences of all eleven species of the Didiereaceae and several outgroup taxa from the Portulacaceae. Results indicated that: 1) the Didiereaceae were embedded within the Portulacaceae, withCalyptrotheca as the sister group of the family; 2) present generic limits were supported; 3)Alluaudiopsis was the most basal lineage; 4) at least two separate episodes of polyploidization within the genusAlluaudia had occurred, and 5) unusually low amounts of variation were present in rapidly evolving noncoding plastid sequences.     

Molecular analyses suggest a need for a significant rearrangement of Rutaceae subfamilies and a minor reassessment of species relationships withinFlindersia

DNA sequencing has been used to construct two molecular phylogenies at the intrafamily and intrageneric level within the Rutaceae. Analysis oftrnL-trnF sequence data for five Rutaceae subfamilies has shown that there is no molecular support for the current subfamily classifications within the Rutaceae. The Dictyolomatoideae and Spathelioideae belong to a clade separate from the clades containing the remaining Rutaceae subfamilies. Rutoideae and Citroideae do not form discrete clades which suggests a reassessment of the subfamily classification is necessary, particularly asRuta falls within the majority Citroideae clade. Flindersioideae forms a clade within the Rutaceae and does not form a separate family or form a clade with Meliaceae.Sequencing of 17Flindersia species produces a similar phylogeny to that proposed by other authors using morphological methods with two exceptions. The molecular phylogeny indicatesF. amboinensis is associated withF. fournieri andF. laevicarpa and, in addition,F. oppositifolia andF. pimenteliana were found to be genetically identical.     

Phylogenetic Use of Noncoding Regions in the GenusGentianaL.: ChloroplasttrnL (UAA) Intron versus Nuclear Ribosomal Internal Transcribed Spacer Sequences

Sequence divergence was estimated within noncoding sequences of both chloroplast DNA (cpDNA)trnL (UAA) intron and nuclear ribosomal DNA (nrDNA) internal transcribed spacer sequences (ITS1 and ITS2) for 10 species of the genusGentianaL. (Gentianaceae). Comparisons of evolutionary rates among these sequences (cpDNA versus nrDNA, ITS1 versus ITS2) were performed. It appears that sequence divergence is on average two to three times higher in ITSs than in thetrnL intron sequences and higher in ITS1 than in ITS2. Both the cpDNA intron and ITSs of nrDNA give concordant phylogenetic trees. However, the ITS-based phylogeny displays higher bootstrap values. At the intrageneric level, at least inGentiana,ITSs (especially ITS2) sequences seem to be more appropriate in the assessment of plant phylogenies. Nevertheless, the cpDNAtrnL intron seems to be preferable at the intergeneric level.     

Phylogenetic Analyses of Teleki Grapevine Rootstocks Using Three Chloroplast DNA Markers

Teleki rootstocks are used in grapevine-producing countries all over the world. They represent one of the largest groups of available rootstocks but their origin is still in dispute although they have been regarded as Vitis berlandieri × V. riparia hybrids. To investigate their possible origin, we amplified and sequenced three chloroplast regions, two non-coding spacers (trnL-F, trnS-G) and the trnL group I intron in a core collection of Teleki rootstocks representing widespread accessions and related wild North American grape species (V. berlandieri, V. riparia and V. rupestris). Concatenated sequence data coupled with microstructural changes discovered in the chloroplast regions provided data to trace the maternal ancestry of the Teleki lines. All chloroplast regions showed both nucleotide and length variation. Length mutations in the non-coding regions represented mostly simple sequence repeats of poly-A and -T stretches. These indel characters exhibited additional diversity comparable with the nucleotide diversity and increased resolution of the phylogenetic trees. We found that a group of Teleki accessions position together with the wild grape species V. riparia. Another group of Teleki rootstocks formed a sister group to the other North American species V. berlandieri. These clades had moderate support values, and they do not share ancestry with other accessions of Teleki rootstocks resolved with high support value in the V. riparia clade. It seems that Teleki-Kober 5BB and 125 AA accessions might have a V. berlandieri maternal background. We also found great differences within putative clones of Teleki 5C and Teleki-Kober 5BB suggesting that the selection of these accessions was performed on heterogenous or mislabeled plant material collectively maintained under these names.     

Evolution of 4-coumarate:coenzyme A ligase (4CL) gene and divergence of Larix (Pinaceae)

The evolutionary dynamics of the 4CL gene encoding 4-coumarate:coenzyme A ligase was investigated in the genus Larix (Pinaceae) by comparing copy number, GC content and codon usage, sequence divergence, and phylogenetic analysis. All 4CL clones of Larix formed a strongly supported monophyletic group, in which two robust clades (4clA and 4clB) derived from an ancient gene duplication event in the common ancestor of Larix were identified. Further gene duplication in the 4clA clade gave rise to two subclades 4clA(1) and 4clA(2). Frequent duplication/deletion appears to be a common evolutionary phenomenon in the 4CL gene family and paralogous genes differ greatly in their evolution rate. The existence of L. speciosa in subclades 4clA(1) and 4clA(2) suggests that this species may represent a primitive form of Larix or the closest relative of the common ancestor of the Eurasian Sect. Multiserialis. In addition, cpDNA and nrDNA ITS analyses support the hypothesis of an early separation of Larix into a North American and a Eurasian clade, which is congruent with the results of previous allozyme and very recent AFLP analyses. The unexpected close relationship between North American larches and the short-bracted species L. gmelinii in East Asia, based on the 4CL gene tree, may stem from lineage sorting.     

Genetic evidence and species distribution modelling reveal the response of Larix sibirica and its related species to Quaternary climatic and ancient historical events

Larix Mill., a boreal forest genus, distributes in Eurasia and North America. Larix sibirica and some related species of this genus have wide distributions in the northeastern, northwestern and southwestern regions of China. To elucidate the response of populations to different regional historical events, we investigated the spatial genetic structures and the divergence time of genotypes in these Larix species based on two cpDNA fragments. To approach this question, we have also modelled the change of glacial-interglacial distributions for populations in different regions using the species distribution modelling. The results of analyses of molecular variance indicate genetic variation mainly occurred between three isolated regions (74.4%). Distributions of Larix have possibly undergone contractions during the Last Glacial Maximum (LGM) period in comparison with their present-day distributions in all three regions. Recent rapid uplift of Qinghai-Tibet Plateau likely promoted the high level of genetic diversity (π = 0.69 × 10⁻³, Hd = 0.711) and Plio-Pleistocene (1.9–2.7 Ma) lineage divergence in the southwestern region. Founder effect and genetic drift accompanied by postglacial colonizations are expected to shape one dominated haplotype and southern LGM distribution in the northeastern region. Quaternary Glacial period is also speculated to trigger low level of genetic diversity (π = 0.09 × 10⁻³, Hd = 0.133) in the northwestern region. Disjunction of L. sibirica and lineage divergence at about 1.6 Ma in the Altai Mountains and the eastern Tianshan Mountains are related to the enhanced aridity in the Asian interior and the desert expansion in Dzungarian Basin during the early Pleistocene.