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!"#$%&2020 40 ): 0969-0977 犃犮狋犪犅狅狋 犅狅狉犲犪犾 犗犮犮犻犱犲狀狋 犛犻狀 犱狅犻 10.7606 j.issn.10004025.2020.06.0969 http :// xbzwxb.allj ournal.net !"#$20200209 %&"!'#$20200509 ()*+'!()*+%,-./012019JM476 ); !23+401562020KJWL21 ,-./7 81983- ), 9:;<=>?@A#$BCDE%./Email jinglee211 163.com FGHIJKK:;LMNO;./DP=>?@A#$QRD$SH./Email missilexll 163.com (0 犐犛犛犚 123456789:; <=>?@1A ! " #$% &'( )*+ ,-- !2TU%V D$QWXYZ%V#$RD$[\./]^_`a!2 710065 !23bc4defgh!2 710061 B C!ijk狔狆 狉犻 犲犱犻狌犿狋犻犫犲狋犻犮狌犿 lmnopqrstuvwxk+#$yz{|g}~x #$./uISSR BC4d)g}!x !ijk B|g}D!ijkxU ¡¢£¤¥:( @100¦ISSR §$g¨©ª«12¦ Ep]¬-x§$¨®¯«136°^E°^±B²犘犘犅 ³100% ´µ Neis,¶·¸s ShannonG¹·¸Bº|0.3186s0.4843 »x Neis¼½¾0.0333 0.1701¿»Neis ÀÁ¾0.8435 0.9673¿»´µBÃĸst s,¶ÅBº|0.2229s1.7430 。( UP GMA ÇsÈÉÇxÄÊẸ̈ͤ¥«ÎÏQ'!¿»Ð¾BôµÑ!ijk» BÃQUÒ½¿»ÀÓÔ¤Õ./Ö|!ijk¾BC×ØÑmnÙÚxÛÎÏ Q'!yÜDÝBÃISSR BCÞuÆ!ijk»Bà ßàáx./DEF!ijkISSR GH1IJQ346 .5 Q789 KL3MN犌犲狀犲狋犻犮犇犻狏犲狉狊犻狋 狅犳 狔狆 狉犻 犲犱犻狌犿狋犻犫犲狋犻犮狌犿 犘狅 狌犾犪狋犻狅狀狊犚犲狏犲犪犾犲犱犫 犐犛犛犚犃狀犪犾 狊犻狊 LIJing ZHANGXiaofei CHENJueyi WU Linqin XULingling 1 Key LaboratoryofPlantmicrobeCollaboration Research SchoolofBiologicaland EnvironmentalEngineering Xian University Xian710065 China 2XianAgriculturalTechnologyExtensionCentre Xian710061 China 犃犫狊狋狉犪犮狋 狔狆 狉犻 犲犱犻狌犿狋犻犫犲狋犻犮狌犿isauniqueorchidspecieswithhighornamentalandmedicinalvalue.As oneendangeredspecies itiscurrentlylistedasanationalprotectedplant.Inthisstudy geneticstructure andgeneticdiversityof7 狋犻犫犲狋犻犮狌犿populationscollectedinwesternChinawasanalyzedusingISSRmo lecularmarkers.Thisstudycanprovideareferenceforspeciesconservationinnaturalhabitatsof 狋犻犫犲狋犻 犮狌犿.Theresultsshowed :( 12polymorphicISSR markerswithstrongrepeatabilitywerechosenfrom 100ISSRprimers.Atotalof136bandswereidentified andthepercentageofpolymorphicbands 犘犘犅 was100%.Neisgenediversity andShannonsinformationindex were0.3186and0.4843 re spectively.TheNeisgeneticdistancerangedfrom0.0333to0.1701andgeneticsimilarityrangedfrom

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Page 1: (0犐犛犛犚 123456789:; ?@1A

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,2020,40(6):0969-0977犃犮狋犪犅狅狋.犅狅狉犲犪犾.犗犮犮犻犱犲狀狋.犛犻狀.

  犱狅犻:10.7606/j.issn.10004025.2020.06.0969                   http://xbzwxb.alljournal.net

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犌犲狀犲狋犻犮犇犻狏犲狉狊犻狋狔狅犳犆狔狆狉犻狆犲犱犻狌犿狋犻犫犲狋犻犮狌犿

犘狅狆狌犾犪狋犻狅狀狊犚犲狏犲犪犾犲犱犫狔犐犛犛犚犃狀犪犾狔狊犻狊

LIJing1,ZHANGXiaofei2,CHENJueyi

1,WULinqin1,XULingling

(1KeyLaboratoryofPlantmicrobeCollaboration Research,SchoolofBiologicalandEnvironmentalEngineering,Xian

University,Xian710065,China;2XianAgriculturalTechnologyExtensionCentre,Xian710061,China)

犃犫狊狋狉犪犮狋:犆狔狆狉犻狆犲犱犻狌犿狋犻犫犲狋犻犮狌犿isauniqueorchidspecieswithhighornamentalandmedicinalvalue.As

oneendangeredspecies,itiscurrentlylistedasanationalprotectedplant.Inthisstudy,geneticstructure

andgeneticdiversityof7犆.狋犻犫犲狋犻犮狌犿populationscollectedinwesternChinawasanalyzedusingISSRmo

lecularmarkers.Thisstudycanprovideareferenceforspeciesconservationinnaturalhabitatsof犆.狋犻犫犲狋犻

犮狌犿.Theresultsshowed:(1)12polymorphicISSRmarkerswithstrongrepeatabilitywerechosenfrom

100ISSRprimers.Atotalof136bandswereidentified,andthepercentageofpolymorphicbands(犘犘犅)

was100%.Neisgenediversity(犎e)andShannonsinformationindex(犐)were0.3186and0.4843,re

spectively.TheNeisgeneticdistancerangedfrom0.0333to0.1701andgeneticsimilarityrangedfrom

Page 2: (0犐犛犛犚 123456789:; ?@1A

0.8435to0.9673amongpopulations.Theoverallgenedifferentiationcoefficient(犌st)was0.2229and

theestimatedvalueofgeneflowwas1.7430.(2)TheUPGMAandNeighborJoiningclusteringanalysis

bothindicatedcleardifferentiationbetweenpopulationsfromSichuanandShaanxi.Theoverallcorrelation

betweengeneticdistanceandgeographicaldistancewasnotsignificant.Thisstudyindicatedhighgenetic

diversityin犆.狋犻犫犲狋犻犮狌犿populationsandgreatdivergencebetweenSichuanandShaanxipopulationsonmo

lecularlevel.ISSRmarkerscouldbeusedforinvestigatingthegeneticstructure,diversityandgeneticdif

ferentiationof犆.狋犻犫犲狋犻犮狌犿populations.

犓犲狔狑狅狉犱狊:犆狔狆狉犻狆犲犱犻狌犿狋犻犫犲狋犻犮狌犿;geneticdiversity;geneticstructure;ISSR

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S1 6789:;OP?TUVWX

Table1 Collectioninformationof犆狔狆狉犻狆犲犱犻狌犿狋犻犫犲狋犻犮狌犿populationsinthisstudy

��õö

Populationcode

�Ö�^

Location=>

Altitude/m��¸

Samplesize�U÷�

Geocoordinate

�Öç»

Collectiondate(yearmonth)

SP ÎÏøùúSongpanCounty,Sichuan 3190~3365 64 32.75°N,103.82°E 201506

KD ÎÏûñ KangdingCounty,Sichuan 2891~2945 23 30.00°N,101.94°E 201607

XJ ÎÏ%üú XiaojinCounty,Sichuan 3494~3720 26 31.15°N,102.76°E 201706

PW ÎÏØýúPingwuCounty,Sichuan 3385~3410 13 32.58°N,104.62°E 201706

GZ ÎÏþÿú GanziCounty,Sichuan 3780~3826 10 31.53°N,99.74°E 201607

TB '!! ú TaibaiCounty,Shannxi 3066~3219 15 33.98°N,107.75°E 201706

MX '!yú MeixianCounty,Shannxi 3210~3274 12 34.02°N,107.97°E 201706

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ShannonG¹·¸

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(犎t)|0.3097±0.0230,��{,¶���

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(SP)��1~22A�µ

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Z1 §$810(A)、811(B)、834(C)、847(D)¾�B!ijk_`�;gISSRPCRNO�£

A.1-18.SichuanKangding(KD)populationsampleNo.1-18;B,C.1-22.SichuanSongpan(SP)populationsample

No.1-22;D.1-18.SichuanXiaojin(XJ)populationsampleNo.1-18;M.DL2000(bandsizesfromsmalltolargeare100bp,

250bp,500bp,750bp,1000bpand2000bp)

Fig.1 ISSRPCRfingerprintsofpart犆.狋犻犫犲狋犻犮狌犿samplesusingprimer810(A),811(B),834(C)and847(D)

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Page 5: (0犐犛犛犚 123456789:; ?@1A

S2 犐犛犛犚]^WXabcij

Table2 InformationofISSRprimersandamplificationresults

§$�A

Primercode§$�{

Primersequence(5′→3′)3IJÂ

Tm/℃

´¦%¸

Totalnumberofbands

�E�¦%¸

Numberofpolymorphicbands

�E�¦%=x

Percentageofpolymorphicbands(犘犘犅)/%

810 GAGAGAGAGAGAGAGAT 53 16 16 100

811 GAGAGAGAGAGAGAGAC 54 14 14 100

834 AGAGAGAGAGAGAGAGYT 53 15 15 100

835 AGAGAGAGAGAGAGAGYC 51 15 15 100

836 AGAGAGAGAGAGAGAGYA 51 11 11 100

847 CACACACACACACACARC 54 9 9 100

855 ACACACACACACACACYT 55 9 9 100

857 ACACACACACACACACYG 54 13 13 100

876 GATAGATAGACAGACA 49 9 9 100

881 GGGTGGGGTGGGGTG 54 10 10 100

887 DVDTCTCTCTCTCTCTC 59 5 5 100

895 GTTGGTAGCTCTTGATC 55 10 10 100

´0Total 136 136 100

S3 67897l:;犐犛犛犚<=>?@fg

Table3 Thegeneticdiversitystatisticsof7populationsof犆.狋犻犫犲狋犻犮狌犿

��

Population

q¯ß°,¶¸

Observednumberofalleles(犖a)

nhß°,¶¸

Effectivenumberofalleles(犖e)

Neis,¶���

Neisgenediversity(犎e)

ShannonG¹·¸

Shannonsinformationindex(犐)

�E�°^¸

Numberofpolymorphicloci

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Percentageofpolymorphicloci/%

SP 1.9485 1.5007 0.2958 0.4482 129 94.85

KD 1.8309 1.4236 0.2566 0.3931 113 83.09

XJ 1.7941 1.3586 0.2207 0.3433 108 79.41

PW 1.7574 1.4026 0.2401 0.3651 103 75.74

GZ 1.6324 1.3608 0.2125 0.3204 86 63.24

TB 1.7132 1.4024 0.2336 0.3512 97 71.32

MX 1.6176 1.3950 0.2253 0.3331 84 61.76

ØÍw Average 1.7563 1.4062 0.2407 0.3649 102.86 75.63

´�µTotal 2.0000 1.5343 0.3186 0.4843 136 100

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Table4 Neisgeneticidentityandgeneticdistanceamong7populationsof犆.狋犻犫犲狋犻犮狌犿

��Population SP KD XJ PW GZ TB MX

SP — 0.9052 0.8966 0.9011 0.8783 0.8843 0.8767

KD 0.0996 — 0.9196 0.9063 0.8747 0.8777 0.8597

XJ 0.1091 0.0838 — 0.9673 0.9515 0.8905 0.8595

PW 0.1041 0.0984 0.0333 — 0.9272 0.8756 0.8537

GZ 0.1298 0.1339 0.0497 0.0756 — 0.8672 0.8435

TB 0.1230 0.1304 0.1159 0.1328 0.1425 — 0.9621

MX 0.1316 0.1512 0.1514 0.1581 0.1701 0.0387 —

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S5 67897l:;犐犛犛犚34512Yt1A

Table5 Molecularvarianceanalysis(AMOVA)in7populationsof犆.狋犻犫犲狋犻犮狌犿basedonISSRmarkers

ÎÏ|�

Sourceofvariation)ûÂ

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��{ Withinpopulations 158 183.266 1.7842 74.42 <0.001

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GZ.SichuanGanzipopulation;TB.ShannxiTaibaipopulation;

MX.ShannxiMeixianpopulation

Fig.2 Phylogeneticclusteringof7populationsof

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andNeighborJoining(B)method

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