A Research Of The Effects Of Altitude On Hereditary Property Of Elymus L.

Elymus L.sensu lato,including about 150 allopolyploid species distributed in a wide range of ecological habitats over the temperate and subtropic regions,is the largest genus of the tribe Triticeae in the Poaceae family.The St,H and Y genome in various combinations(polyploidization and hybridization)constitute Elymus species.The St genome,which originated from Pseudoroegneria(Nevski)á.L?ve,while H genome originated from Hordeum L.,no putative Y genome diploids have been identified.The Elymus species is also a mainly germplasm resources covering excellent genomes for disease resistance,stress tolerance,wide adaptation,high protein and lysine contents.Thus it provides gene resources for the breeding of wheat crops and improvement of forage quality.It was well known that the Elymus species showed strong adaptability to altitude,and its inheritance and/or variation had great relationship with environment.Did altitude gradient promote the event of differentiation? Available molecular phylogenetic data are needed to investigate this issue.In the present study,tetraploids species(Elymus burchan-buddae,St Y)and hexaploid species(Elymus nutans,St HY)were selected.And their nuclear DNA ITS(ribosomal internal transcribed spacers),Waxy(Granule-bound Starch Synthase Gene)and Adh(alcohol dehydrogenase)genes were cloned,sequenced and statistically analyzed to explore the influence of altitude on genetic characteristics of Elymus L.The selected materials coming from nine different altitudinal gradients were similar in morphological and had overlapping geographical distribution.The results were as follows:1.Seeds of the collected materials were germinated and grown in a green-house for DNA extraction.The total DNA was successfully isolated from fresh leaves through the CTAB procedure.2.By means of selecting appropriate primers,PCR amplification,cloning and sequencing,3 nuclear DNA gene sequences of Elymus nutans and Elymus burchan-buddae were obtained.The sequence of each clone was compared using the BLASTN program,and phylogenetic analyses of three data matrices including Waxy,Adh2 and ITS sequences were carried out using the Maximum-parsimony(MP)method and neighbour-joining(NJ)method.3.A total of 46 ITS sequences from Elymus nutans showed clear distinction between St and H genome.The ITS sequence informative character from St genome was 1.1%,while it was 6.7% from H genome,indicating that ITS sequence informative character of Elymus nutans from different altitudes was not rich enough.The G+C content of ITS sequences from St and H genome didn’t differ much at all,and there was no obvious distinction among ITS sequences.4.A total of 55 Waxy sequences from Elymus nutans showed clear distinctionbetween St and H genome.The Waxy sequence informative character from St genome was 3.3%,while it was 14.0% from H genome,suggesting that Waxy sequence informative character from St genome was not rich enough,while it was relatively rich from H genome.Similarly,the G+C content of Waxy sequences from St and H genome didn’t differ much at all.5.A total of 47 Adh2 sequences from Elymus nutans showed clear distinction between St and H genome.The Adh2 sequence informative character from St genome was 0.6%,and it was 0.24% from H genome,indicating that Adh2 sequence informative character of Elymus nutans from different altitudes was not rich enough.Meanwhile,the G+C content of Adh2 sequences from St and H genome didn’t differ much at all,and there was no obvious difference among Adh2 sequences.6.The analysis of 51 ITS sequences,54 Waxy sequences and 29 Adh2 sequences from Elymus burchan-buddae showed that the ITS,Waxy and Adh2 sequence informative character was 2.1%,6.4% and 0.6%,respectively.All these three genes sequence informative characters of Elymus burchan-buddae from different altitudes were not rich enough.Meanwhile,the G+C content of three nuclear genes among different samples didn’t differ much at all,and there was no obvious distinction among different sequences.7.Both of the Waxy,Adh2 and ITS phylogenetic trees based on MP and NJ analysis revealed that all investigated Elymus nutans materials could be mainly divided into two clades(St and H).In both clades,multiple populations from different altitudes distributed in the same sub branch,while individuals from the same population distributed in distinct sub branch,showing no significant differences among populations.The individual diversity from the same population was even greater than that of among populations.At the same time,bootstrap values of different sub branch were low,and it is likely that the surveyed materials belonged to the same species while without much phylogenetic information and obvious differentiation.In general,it could be concluded that the genetic diversity of Elymus nutans did not vary depending on altitudes.8.The Waxy,Adh2 and ITS phylogenetic trees showed that all investigated Elymus burchan-buddae materials only contained the Waxy,Adh2 and ITS sequences from St genome.In each branch,multiple populations from different altitudes distributed in the same sub branch,while individuals from the same population distributed in distinct sub branch,showing no significant differences among populations.The individual diversity from the same population was even greater than that of among populations.In addition,bootstrap values of different sub branch were low,and it is likely that the surveyed materials belonged to the same species while without much phylogenetic information and obvious differentiation.In brief,it could be concluded that the genetic diversity of Elymus burchan-buddae did not vary depending on altitudes.9.According to part of the phylogenetic trees based on three nuclear genes from Elymus nutans and Elymus burchan-buddae materials,we could see that there was a close genetic distance between high altitude and low altitude populations.Besides,close genetic distance occurred within populations at intermediate altitudes.Thissuggested that there was certain regularity in genetic diversity along with altitudes,and further research is needed to get better insights into this issue.10.In phylogenetic tree based on Waxy sequences from Elymus burchan-buddae materials,there was a sub branch consisting of sequences with a length of 1367 bp.It was related to Waxy sequence from St genome through comparison using the BLASTN program,and it probably came from Y genome.Further research is also needed to prove this viewpoint.