هﺪﮑﺸﻧاد
ي يزروﺎﺸﮐ مﻮﻠﻋ
ﯽﺷزﻮﻣآ هوﺮﮔ تﺎﺗﺎﺒﻧ حﻼﺻا و ﺖﻋارز
نﺎﯾﺎﭘ ﻪﻣﺎﻧ ﻪﺟرد ﺖﻓﺎﯾرد ياﺮﺑ ﺪﺷرا ﯽﺳﺎﻨﺷرﺎﮐ ي
ﻪﺘﺷر رد ي
يزروﺎﺸﮐ ﯽﺳﺪﻨﻬﻣ ﺶﯾاﺮﮔ
يزروﺎﺸﮐ رد يژﻮﻟﻮﻨﮑﺗﻮﯿﺑ
:ناﻮﻨﻋ
يراﺬﮔﺪﮐرﺎﺑ ﯽﯾوراد نﺎﻫﺎﯿﮔ زا ﯽﺧﺮﺑ DNA
ﻪﺑ ﻞﯿﺑدرا ﯽﻣﻮﺑ يزﺎﺳﺪﻨﺘﺴﻣ رﻮﻈﻨﻣ
نآ ﯽﺘﺴﯾز عﻮﻨﺗ ﺎﻫ
دﺎﺘﺳا (ﺪﯿﺗﺎﺳا) ﺎﻤﻨﻫار
:
ﺮﺘﮐد هداﺰﻧﺎﻣﺮﻬﻗ بﺎﺑر ﺮﺘﮐد ،نﺎﺘﺳژد ارﺎﺳ
دﺎﺘﺳا (ﺪﯿﺗﺎﺳا) روﺎﺸﻣ
:
ﺮﺘﮐد لآ ﯽﻘﺗ ﺪﻤﺤﻣ ﺮﺘﮐد ،ﻮﺠﻣزر ﻞﯾاﺮﺒﺟ ﻢﯿﻫاﺮﺑا
ﺮﮕﺸﻫوﮋﭘ :
يﺪﺳا ﻪﻤﻃﺎﻓ
رﺎﻬﺑ - 1394
مﺎﻧ ﯽﮔداﻮﻧﺎﺧ ﻮﺠﺸﻧاد
ا : يﺪﺳ مﺎﻧ
ﻪﻤﻃﺎﻓ :
ناﻮﻨﻋ نﺎﯾﺎﭘ ﻪﻣﺎﻧ :
يراﺬﮔﺪﮐرﺎﺑ ﻪﺑ ﻞﯿﺑدرا ﯽﻣﻮﺑ ﯽﯾوراد نﺎﻫﺎﯿﮔ زا ﯽﺧﺮﺑDNA
نآ ﯽﺘﺴﯾز عﻮﻨﺗ يزﺎﺳﺪﻨﺘﺴﻣ رﻮﻈﻨﻣ ﺎﻫ
دﺎﺘﺳا (ﺪﯿﺗﺎﺳا) ﺎﻤﻨﻫار ﺮﺘﮐد: هداﺰﻧﺎﻣﺮﻬﻗ بﺎﺑر ﺮﺘﮐد ،نﺎﺘﺳژد ارﺎﺳ
دﺎﺘﺳا (ﺪﯿﺗﺎﺳا) وﺎﺸﻣ ﺮﺘﮐد :ر لآ ﯽﻘﺗ ﺪﻤﺤﻣ ﺮﺘﮐد ،ﻮﺠﻣزر ﻞﯾاﺮﺒﺟ ﻢﯿﻫاﺮﺑا
ﻊﻄﻘﻣ ﻠﯿﺼﺤﺗ ﺪﺷرا ﯽﺳﺎﻨﺷرﺎﮐ:ﯽ
ﻪﺘﺷر يزروﺎﺸﮐ ﯽﺳﺪﻨﻬﻣ :
ﺶﯾاﺮﮔ يزروﺎﺸﮐ رد يژﻮﻟﻮﻨﮑﺗﻮﯿﺑ :
هﺎﮕﺸﻧاد ﻖﻘﺤﻣ: ﻠﯿﺑدرا ﯽ
هﺪﮑﺸﻧاد يزروﺎﺸﮐ مﻮﻠﻋ:
ﺦﯾرﺎﺗ :عﺎﻓد /27 /3 94
داﺪﻌﺗ ﺤﻔﺻ :تﺎ 99
هﺪﯿﮑﭼ :
ا ﺎﻔﺘﺳ هد ﺢﯿﺤﺻ زا ﺎﻫﺎﯿﮔ ن وراد ﯾـ ﯽ ﺪﻨﻣزﺎﯿﻧ ﺑ ـ ﻪ و ﺟ ـﻮ د ا ﺎﻋﻼﻃ ت د ﻖﯿﻗ و ﯽﻠﻤﻋ ا ﺳ .ت ﺪﮐرﺎﺑ ﻦﻓ يراﺬﮔ سﺎﺳاﺮﺑDNA
تادﻮﺟﻮﻣ ﻦﯿﺑ يﺪﻧوﺎﺸﯾﻮﺧ ﻂﺑاور يزﺎﺳزﺎﺑ و ﻪﻌﻟﺎﻄﻣ هدر ﻒﻠﺘﺨﻣ حﻮﻄﺳ رد هﺪﻧز
و ﻪﺘﺳار ،هداﻮﻧﺎﺧ ،ﺲﻨﺟ ،ﻪﻧﻮﮔ) يﺪﻨﺑ
(...
هداد زا هدﺎﻔﺘﺳا ﺎﺑ ﯽﻣ ﯽﻟاﻮﺗ يﺎﻫ
ﺪﺷﺎﺑ ﻦﯾا دﻮﺟوﺎﺑ ، نژ داﺪﻌﺗ ﺶﯾاﺰﻓا ﺎﺑ
ﻊﻗاو ﻢﻬﻓ ،ﻪﻌﻟﺎﻄﻣ درﻮﻣ يﺎﻫ ﻪﻧﺎﻨﯿﺑ
ﻂﺑاور زا يﺮﺗ
هدر ﻒﻠﺘﺨﻣ حﻮﻄﺳ رد يﺪﻧوﺎﺸﯾﻮﺧ ﯽﻣ ﻞﺻﺎﺣ يﺪﻨﺑ
.ددﺮﮔ ﻦﯾا ﻦﻓ ياﺮﺑ ﯽﯾﺎﺳﺎﻨﺷ هد
وراد هﺎﯿﮔ ﯽﯾ ،ﻪﻧﻮﺑﺎﺑ ،ﺐﻠﻌﺛ)
ﻦﺳﻮﺳ ،ﺮﯿﺸﮐﺎﺧ ﻦﯾﺮﯿﺷ ،ﻪﻨﻣرد ،ﯽﻨﺳﺎﮐ ،غاﺮﭽﻠﭼ
ﻪﻧﻮﭘ ،نﺎﯿﺑ (ناردﺎﻣﻮﺑ ،ﮓﻨﻫرﺎﺑ ،ﺎﺳ
ﻊﻤﺟ يروآ ﺷ ﻞﯿﺑدرا نﺎﺘﺳا زا هﺪ
ﺪﺷ هدﺎﻔﺘﺳا جاﺮﺨﺘﺳا .
شور ﻪﺑ DNA ﺮﯿﯿﻐﺗ ﻪﺘﻓﺎﯾ و ﺖﻓﺮﮔ مﺎﺠﻧا CTAB
ﺎﺑ PCR ﯽﺣاﺮﻃ يﺎﻫﺮﮔزﺎﻏآ ﺮﺑ هﺪﺷ
سﺎﺳا
يﺎﻫﺪﮐرﺎﺑ ﯽﺘﺳﻼﭘوﺮﻠﮐ
،rbcL trnH-psbA ،
ﻪﺘﺴﻫ ﺪﮐرﺎﺑ وmatK يا
ﺲﭙﺳ .ﺪﺷ مﺎﺠﻧاITS لﻮﺼﺤﻣ
ﺺﻟﺎﺧPCR يزﺎﺳ
.ﺪﯾدﺮﮔ ﯽﻟاﻮﺗ ﻦﯿﯿﻌﺗ و هﺪﺷ ﺪﺻرد
ﺖﯿﻘﻓﻮﻣ ﯽﻟاﻮﺗ و ﺮﯿﺜﮑﺗ ﺎﯾ
ﻪﻧﻮﻤﻧ رد ﺎﻫﺪﮐرﺎﺑ ﯽﺑ ﯽﺳرﺮﺑ درﻮﻣ يﺎﻫ
ﻪﺑ ﺐﯿﺗﺮﺗ و70
،50
و60 ،40 و50 ،10 و70 ﻪﺑ40 ﺖﺳد ﺪﻣآ . ﻪﺑ ﻧﻮﮔ ﮏﯿﮑﻔﺗ رد ﺪﮐرﺎﺑ نﺎﮑﻣ ﺮﻫ ﯽﯾﺎﻧاﻮﺗ ﺢﻄﺳ ﯽﺑﺎﯾزرا رﻮﻈﻨﻣ ﻪ
ﯽﻫﺎﯿﮔ يﺎﻫ
هداد تﺎﻋﻼﻃا هﺎﮕﯾﺎﭘ زا ﺲﻨﺟ نﺎﻤﻫ ﻪﺑ ﻖﻠﻌﺘﻣ ﺮﮕﯾد ﯽﻟاﻮﺗ ﺪﻨﭼ ﻪﻌﻟﺎﻄﻣ درﻮﻣ ﻊﻤﺟNCBI
.ﺪﺷ يروآ ﯽﻟاﻮﺗ
ﻪﻧﻮﻤﻧ ﺎﺑ ﺎﻫ -
يﺎﻫ دﻮﺟﻮﻣ رد هداد هﺎﮕﯾﺎﭘ ﻒﯾدﺮﻤﻫNCBI
ﺖﺤﺗ و هﺪﺷ ﯽﮑﯿﺗﺎﻣرﻮﻔﻧاﻮﯿﺑ ﻪﯾﺰﺠﺗ
ﺪﻨﺘﻓﺮﮔ راﺮﻗ .
رد يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد
ﻪﻧﻮﮔ ﻢﻫ يﺎﻫ ﺮﺑ ﺲﻨﺟ ﯽﻟاﻮﺗ سﺎﺳا ﺪﮐرﺎﺑ يﺎﻫ
ﻫ يﺎ وrbcL trnH-psbA رد .ﺪﻧﺪﺷ ﮏﯿﮑﻔﺗ ﻢﻫ زا
ﺪﮐرﺎﺑ ﻂﻘﻓ ﺮﻣا ﻦﯾاITS
رد ﻦﯾﺮﯿﺷ هﺎﯿﮔ نﺎﯿﺑ
هﺎﯿﮔ ،ﻪﻌﻟﺎﻄﻣ ﻦﯾا رد .دﻮﺑ قدﺎﺻ ﻦﺳﻮﺳ
ﺎﺑ رﺎﺑ ﻦﯿﻟوا ياﺮﺑ غاﺮﭽﻠﭼ ﺪﮐرﺎﺑ
ﺪﺷ يراﺬﮔﺪﮐرﺎﺑrbcL .
ﺪﮐرﺎﺑSNP يﺎﻫ ﻢﮐ)rbcL زا ﺮﺗ
،(30 trnH-psbA ﻢﮐ)
زا ﺮﺗ (100 و ﻢﮐ)matK زا ﺮﺗ (20 ﺪﺷ شرﺎﻤﺷ .
ﺎﻣا ﺪﮐرﺎﺑSNP
ﻦﯾﺮﯿﺷ رد ﺎﻬﻨﺗITS ﻪﻧﻮﻤﻧ ﺮﮕﯾد ياﺮﺑ و دﻮﺑ لﻮﺒﻗ ﻞﺑﺎﻗ نﺎﯿﺑ
ﺎﻫ ﺪﮐرﺎﺑ ﻦﯾا ي زا ﺮﺗﻻﺎﺑ
ﻪﻠﺻﺎﻓ ﺪﺻرد .ﺪﺷ شرﺎﻤﺷ دﺪﻋ200
ﯽﻟاﻮﺗ ﺎﺑ ﻪﻌﻟﺎﻄﻣ درﻮﻣ نﺎﻫﺎﯿﮔ ﻦﯿﺑ ﯽﮑﯿﺘﻧژ ﻪﻧﻮﮔ يﺎﻫ
نﺎﻤﻫ زا يﺎﻫ رد ﺪﮐرﺎﺑ و ﺲﻨﺟ
هداد هﺎﮕﯾﺎﭘ ﺪﮐرﺎﺑ ﻪﺳ ردNCBI
،rbcL وmatK trnH-psbA ﻦﯿﯾﺎﭘ
ﺪﺷ دروآﺮﺑ ﺎﺑ ﻪﻌﻟﺎﻄﻣ درﻮﻣ نﺎﻫﺎﯿﮔ ﺪﮐرﺎﺑ ﯽﻟاﻮﺗ يﻻﺎﺑ ﺖﻫﺎﺒﺷ زا نﺎﺸﻧ ﺮﻣا ﻦﯾا ﻪﮐ
ﯽﻟاﻮﺗ دراد ار ﺪﮐرﺎﺑ و ﺲﻨﺟ نﺎﻤﻫ زا هداد هﺎﮕﯾﺎﭘ رد دﻮﺟﻮﻣ يﺎﻫ
،ﻦﯾا دﻮﺟو ﺎﺑ . ﺪﮐرﺎﺑ رد ﺪﺻرد ﻦﯾا
ﺲﻨﺟ رد ﺎﻬﻨﺗITS
ﻦﯾﺮﯿﺷ .ﺪﯾدﺮﮔ دروآﺮﺑ ﯽﻨﯿﯾﺎﭘ راﺪﻘﻣ نﺎﯿﺑ ﺪﮐرﺎﺑ ،ﻦﯾاﺮﺑﺎﻨﺑ
ﻪﺑrbcL ﺗ ترﺪﻗ ﻞﯿﻟد داﺪﻌﺗ ،ﻻﺎﺑ ﮏﯿﮑﻔ
و ﻦﯿﯾﺎﭘSNP ﺖﯿﻌﻣﺎﺟ
ﻪﻧﻮﮔ ﺮﺜﮐا رد ﺎﻫ
، ﻪﺑ ﯽﻓﺮﻌﻣ ﺪﮐرﺎﺑ ﻦﯾﺮﺘﻬﺑ ناﻮﻨﻋ ﺪﺷ
. يﺎﻫﺪﮐرﺎﺑ ،ﻦﯾا دﻮﺟو ﺎﺑ وITS
trnH-psbA ﻪﺑ
ﻞﯿﻟد ﻞﮑﺸﻣ ﻂﺒﺗﺮﻣ
ﺎﺑ ﯽﻟاﻮﺗ لﻮﺼﺤﻣ ﻢﯿﻘﺘﺴﻣ ﯽﺑﺎﯾ مﺪﻋ و PCR
ﺖﯿﻔﯿﮐ ﺎﺑ ﯽﻟاﻮﺗ ﻪﺑ ﯽﺳﺮﺘﺳد
، ﻪﺑ ناﻮﻨﻋ ﻞﻤﮑﻣ يﺎﻫﺪﮐرﺎﺑ .ﺪﻧﺪﺷ ﯽﯾﺎﺳﺎﻨﺷ
ﺪﮐرﺎﺑ ﺑmatK ﻪ ﯽﻟاﻮﺗ و ﺮﯿﺜﮑﺗ ترﺪﻗ ﻞﯿﻟد ﻪﻧﻮﻤﻧ ياﺮﺑ ﻦﯿﯾﺎﭘ ﯽﺑﺎﯾ
ﯽﻤﻧ ﻪﯿﺻﻮﺗ ﯽﺳرﺮﺑ درﻮﻣ يﺎﻫ .دﻮﺷ
ﺪﯿﻠﮐ هژاو ﺎﻫ : نﺎﻫﺎﯿﮔ ﺑ ،ﯽﯾوراد يراﺬﮔﺪﮐرﺎ
، DNA
ﺪﮐرﺎﺑ يﺎﻫ ،rbcL trnH-psbA ،
وmatK ITS
أ
ﺐﻟﺎﻄﻣ ﺖﺳﺮﻬﻓ
ﺐﻟﺎﻄﻣ ناﻮﻨﻋ و هرﺎﻤﺷ
ﻪﺤﻔﺻ
ﻪﻣﺪﻘﻣ :لوا ﻞﺼﻓ
-1 ﻪﻣﺪﻘﻣ1
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. 2
ﺶﻫوﮋﭘ تﺎﯿﻠﮐ :مود ﻞﺼﻓ
-2 ز عﻮﻨﺗ1 ﯽﺘﺴﯾ
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. 6
-2 -1 ﯽﺘﺴﯾز عﻮﻨﺗ ﺖﯿﻤﻫا1
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7
-2 -1 ﺶﯾﺎﺳﺮﻓ2 ﯽﺘﺴﯾز عﻮﻨﺗ
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-2 ﯽﮑﯿﺘﻧژ عﻮﻨﺗ2
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9
-2 ﯽﯾوراد نﺎﻫﺎﯿﮔ3
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. 10
-2 -3 ﺖﯿﻟﻮﺑﺎﺘﻣ عاﻮﻧا1 ﯽﻫﺎﯿﮔ يﺎﻫ
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-2 -3 ﯽﯾوراد نﺎﻫﺎﯿﮔ ﺖﯿﻤﻫا2
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-2 -3 ﺗ هﺎﯿﮔ زا3 وراد ﺎ
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-2 ﺳﻮﺳ4 غاﺮﭽﻠﭼ ﻦ
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. 13
-2 ناردﺎﻣﻮﺑ5
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-2 ﻦﯾﺮﯿﺷ6 نﺎﯿﺑ
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-2 ﻪﻧﻮﺑﺎﺑ7
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-2 ﻪﻨﻣرد8
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-2 ﮓﻨﻫرﺎﺑ9
ب
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-2 ﯽﻨﺳﺎﮐ10
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-2 ﺮﯿﺸﮐﺎﺧ11
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-2 ﻧﻮﭘ12 ﻪ ﺎﺳ
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-2 ﺐﻠﻌﺛ13
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-2 ﯽﯾوراد نﺎﻫﺎﯿﮔ يژﻮﻟﻮﻨﮑﺗﻮﯿﺑ14
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-2 شور15 ﯽﯾوراد نﺎﻫﺎﯿﮔ ﯽﯾﺎﺳﺎﻨﺷ يﺎﻫ
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-2 -15 1 RFLP
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-2 -15 ﺖﺸﮕﻧا2 يرﺎﮕﻧ DNA
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-2 -15 ﺮﮕﻧﺎﺸﻧ3 RAPD
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-2 -15 4 PCR-SSCP
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-2 -15 5 PCR-DGGE
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-2 -15 ﺰﯾر6 هراﻮﻫﺎﻣ ﺎﻫ
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-2 -15 ﻪﯾارآﺰﯾر7 ﺎﻫ
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-2 -15 يراﺬﮔﺪﮐرﺎﺑ8 DNA
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-2 -15 تﺎﺷ يراﺬﮔﺪﮐرﺎﺑ9 نﺎﮔ
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-2 -15 10 AFLP
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-2 ﮏﯿﺗﺎﻣرﻮﻔﻧاﻮﯿﺑ16
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-2 يراﺬﮔﺪﮐرﺎﺑ17 DNA
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ج
-2 -17 ﺪﮐرﺎﺑ شور ﯽﮔﮋﯾو1 يراﺬﮔ
DNA
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-2 -17 ﯽﻟاﻮﺗ2 ﺎﯾ ﯽﺑ
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-2 -17
ﺐﺳﺎﻨﻣ ﺪﮐرﺎﺑ ﮏﯾ ﯽﮔﮋﯾو3
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-2 ﺗ ﺮﺑ يروﺮﻣ18 درﻮﻣ ﯽﻟاﻮ
ﺑ هدﺎﻔﺘﺳا ﻪ ﺪﮐرﺎﺑ ناﻮﻨﻋ
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-2 -18 يراﺬﮔﺪﮐرﺎﺑ1
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-2 -18 نارﻮﻧﺎﺟ رد يراﺬﮔﺪﮐرﺎﺑ2
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-2 -18 -2 چرﺎﻗ رد يراﺬﮔﺪﮐرﺎﺑ1 ﺎﻫ
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36
-2 -18 -2
تاﺮﺸﺣ رد يراﺬﮔﺪﮐرﺎﺑ2
...
...
...
...
37
-2 -18 نﺎﻫﺎﯿﮔ رد يراﺬﮔﺪﮐرﺎﺑ3
...
...
. ...
37
-2 -18 -3 ﯽﻫﺎﯿﮔ يﺎﻫﺪﮐرﺎﺑ1
...
...
...
38
-2 -18 -3 -1
ﺐﺳﺎﻨﻣ ﺪﮐرﺎﺑ ﮏﯾ ﻦﺘﻓﺎﯾ ياﺮﺑ ﻮﺠﺘﺴﺟ1
...
...
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38
-2 -18 -3 -1 يرﺪﻨﮐﻮﺘﯿﻣ مﻮﻧژ2
...
...
...
. 40
-2 -18 -3 -1 ﺖﺳﻼﭘوﺮﻠﮐ مﻮﻧژ3
...
...
...
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41
-2 -18 -3 -1 4 rbcL
...
...
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42
-2 -18 -3 -1 5 matK
...
...
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44
-2 -18 -3 -1 6 trnH-psbA
...
...
...
...
...
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45
-2 -18 -3 -1 7 atpF-atpH و
psbK-psbI
...
46
-2 -18 -3 -1 نوﺮﺘﻨﯾا8 ﻦﯿﺑ يﺎﻀﻓ وtrnL
ﯽﻧژ trnL – trnF
...
...
. ...
...
46
-2 -18 -3 -1 ﺮﮕﯾد يﺎﻫﺪﮐرﺎﺑ9
د
...
..
...
...
....
47
-2 -18 -3 -1 ﻪﺘﺴﻫ مﻮﻧژ10 يا
ITS
....
...
...
48
-2 -18
يراﺬﮔﺪﮐرﺎﺑ رد دﻮﺟﻮﻣ تﺎﻈﺣﻼﻣ4
...
...
51
-2 ﮏﻧﺎﺑ19 ﯽﺗﺎﻋﻼﻃا يﺎﻫ
...
...
...
52
-2 -19 1 GenBank و
NCBI
...
...
....
...
52
-2 -19 2 CBOL
...
...
..
...
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52
-2 -19 3 BOLD
...
...
...
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53
-2
ﺶﻫوﮋﭘ فاﺪﻫا 20
...
....
...
...
54
شور و داﻮﻣ :مﻮﺳ ﻞﺼﻓ ﺎﻫ
-3
يراﺬﮔﺪﮐرﺎﺑ1 DNA
...
...
...
...
...
56
-3 -1 ﻊﻤﺟ1 ﻪﻧﻮﻤﻧ يروآ ﺎﻫ
...
...
...
...
56
-3 -1 جاﺮﺨﺘﺳا2 DNA
...
...
...
56
-3 -1
ﯽﻤﮐ و ﯽﻔﯿﮐ ﯽﺳرﺮﺑ3
...
...
...
...
58
-3 -1 -3 زرﻮﻓوﺮﺘﮑﻟا1
...
...
...
....
58
-3 -1 -3 ﺘﮑﭙﺳا2 ﺮ ﻓو ﺮﺘﻣﻮﺘ
. ...
...
...
...
....
..
59
-3 -1 درﻮﻣ يﺎﻫﺪﮐرﺎﺑ4 هدﺎﻔﺘﺳا
...
...
...
...
...
59
-3 -1 ﺶﻨﮐاو ﻂﯾاﺮﺷ5 PCR
...
...
...
...
59
-3 -1 ﯽﻟاﻮﺗ6 ﻪﻧﻮﻤﻧ ﯽﺑﺎﯾ ﺮﯿﺜﮑﺗ يﺎﻫ
هﺪﺷ
...
..
...
...
...
...
60
-3 -1 ﻪﯾﺰﺠﺗ7 ﯽﻟاﻮﺗ ﺎﻫ
...
...
...
...
60
ه
ﺞﯾﺎﺘﻧ :رﺎﻬﭼ ﻞﺼﻓ
-4 يراﺬﮔﺪﮐرﺎﺑ1 DNA
...
...
...
...
63
-4 -1 درﻮﻣ يﺎﻫﺪﮐرﺎﺑ ﺮﯿﺜﮑﺗ1 ﺎﻄﻣ
ﻪﻌﻟ
...
...
...
...
...
63
-4 -1 درﻮﻣ يﺎﻫﺪﮐرﺎﺑ ياﺮﺑ يراﺬﮔﺪﮐرﺎﺑ ﺞﯾﺎﺘﻧ2 ﻪﻌﻟﺎﻄﻣ
...
...
...
...
...
64
-4 -1 -2 ﺪﮐرﺎﺑ1 rbcL
...
...
...
...
64
-4 -1 -2 ﺪﮐرﺎﺑ2 trnH-psbA
...
...
69
-4 -1 -2 ﺪﮐرﺎﺑ3 matK
...
...
...
72
-4 -1 -2 ﺪﮐرﺎﺑ4 ITS
...
...
...
74
-4 ﻪﺠﯿﺘﻧ 2
تادﺎﻬﻨﺸﯿﭘ و يﺮﯿﮔ
...
...
...
...
77
ﻊﺑﺎﻨﻣ . ...
...
...
...
....
82
لوﺪﺟ ﺖﺳﺮﻬﻓ ﺎﻫ
لوﺪﺟ ناﻮﻨﻋ و هرﺎﻤﺷ
ﻪﺤﻔﺻ
شور و داﻮﻣ :مﻮﺳ ﻞﺼﻓ ﺎﻫ
لوﺪﺟ -3 ﻊﻤﺟ ﻞﺤﻣ :1 ﯽﯾوراد نﺎﻫﺎﯿﮔ يروآ
...
...
...
. 56
وﺪﺟ ل -3 : 2 جاﺮﺨﺘﺳا ﺮﻓﺎﺑ ﻪﯿﻬﺗ هﻮﺤﻧ DNA
...
...
...
..
58
لوﺪﺟ -3 ﺮﮔزﺎﻏآ ﯽﻟاﻮﺗ :3 يﺎﻫ
هدﺎﻔﺘﺳا ﻦﯿﺑ و ﯽﻧژ ﻖﻃﺎﻨﻣ ﺮﯿﺜﮑﺗ ياﺮﺑ هﺪﺷ هدﺎﻔﺘﺳا درﻮﻣ نﺎﻫﺎﯿﮔ ﯽﻧژ
و
...
...
...
...
...
59
ﺞﯾﺎﺘﻧ :مرﺎﻬﭼ ﻞﺼﻓ
لوﺪﺟ -4 و ﺮﯿﺜﮑﺗ رﺎﻣآ :1 ﯽﻟاﻮﺗ
ﻪﻌﻟﺎﻄﻣ درﻮﻣ نﺎﻫﺎﯿﮔ ﯽﺑﺎﯾ
..
...
...
...
...
63
لوﺪﺟ -4 :2 ﺎﻄﻣ درﻮﻣ يﺎﻫﺪﮐرﺎﺑ ﺖﯿﻘﻓﻮﻣ ﺪﺻرد ﻪﻌﻟ
..
...
...
...
...
64
لوﺪﺟ -4 :3 BLASTn ﯽﻟاﻮﺗ
ﺪﮐرﺎﺑ ﯽﺳرﺮﺑ درﻮﻣ يﺎﻫ ﯽﻟاﻮﺗ ﺎﺑrbcL
هداد هﺎﮕﯾﺎﭘ رد دﻮﺟﻮﻣ يﺎﻫ NCBI
...
...
....
...
65
لوﺪﺟ -4 ﯽﮑﯿﺘﻧژ ﻪﻠﺻﺎﻓ :4 ﻪﻧﻮﮔ
ﻪﻧﻮﮔ ﺎﺑ ﯽﺳرﺮﺑ درﻮﻣ يﺎﻫ ﻢﻫ يﺎﻫ
رد دﻮﺧ ﺲﻨﺟ
.NCBI
ﻦﯾﺮﯿﺷ نﺎﯿﺑ )آ ( ﻦﺳﻮﺳ ، غاﺮﭽﻠﭼ
،(ب) ﻪﻧﻮﺑﺎﺑ (ه)ﺮﯿﺸﮐﺎﺧ و (د) ﺐﻠﻌﺛ ،(ج)
. ...
...
...
. ...
67
لوﺪﺟ -4 : 5 BLASTn ﯽﻟاﻮﺗ
يﺎﻫ درﻮﻣ ﯽﺳرﺮﺑ ﺪﮐرﺎﺑ trnH-psbA ﺎﺑ
ﯽﻟاﻮﺗ يﺎﻫ دﻮﺟﻮﻣ رد هﺎﮕﯾﺎﭘ هداد NCBI
...
...
70 لوﺪﺟ -4 ﻪﻧﻮﮔ ﯽﮑﯿﺘﻧژ ﻪﻠﺻﺎﻓ :6 ﻪﻧﻮﮔ ﺎﺑ ﯽﺳرﺮﺑ درﻮﻣ يﺎﻫ
ﻢﻫ يﺎﻫ رد دﻮﺧ ﺲﻨﺟ
. NCBI
ﻪﻧﻮﺑﺎﺑ ) ﮓﻨﻫرﺎﺑ ،(آ) ب
ﺮﯿﺸﮐﺎﺧ و (
) ج (
. ...
...
...
...
...
...
...
...
71
لوﺪﺟ -4 : 7 BLASTn ﯽﻟاﻮﺗ
يﺎﻫ درﻮﻣ ﯽﺳرﺮﺑ ﺪﮐرﺎﺑ ﺎﺑmatK ﯽﻟاﻮﺗ
يﺎﻫ دﻮﺟﻮﻣ رد هﺎﮕﯾﺎﭘ هداد NCBI
...
...
....
73
لوﺪﺟ -4 ﻪﻧﻮﮔ ﯽﮑﯿﺘﻧژ ﻪﻠﺻﺎﻓ :8 ﻪﻧﻮﮔ ﺎﺑ ﯽﺳرﺮﺑ درﻮﻣ يﺎﻫ
ﻢﻫ يﺎﻫ رد دﻮﺧ ﺲﻨﺟ
...NCBI ...
....
73
لوﺪﺟ -4 : 9 BLASTn ﯽﻟاﻮﺗ
يﺎﻫ درﻮﻣ ﯽﺳرﺮﺑ ﺪﮐرﺎﺑ ﺎﺑITS ﯽﻟاﻮﺗ يﺎﻫ دﻮﺟﻮﻣ رد هﺎﮕﯾﺎﭘ هداد NCBI
..
...
...
74
وﺪﺟ ل -4 ﻪﻧﻮﮔ ﯽﮑﯿﺘﻧژ ﻪﻠﺻﺎﻓ :10 ﻪﻧﻮﮔ ﺎﺑ ﯽﺳرﺮﺑ درﻮﻣ يﺎﻫ
ﻢﻫ يﺎﻫ رد دﻮﺧ ﺲﻨﺟ
. NCBI
ﻦﯾﺮﯿﺷ نﺎﯿﺑ
،(آ) ) ﻪﻧﻮﺑﺎﺑ ب ( و
) ﺮﯿﺸﮐﺎﺧ ج (
ز
. ...
...
....
...
...
...
76
ﻞﮑﺷ ﺖﺳﺮﻬﻓ ﺎﻫ
ﻞﮑﺷ ناﻮﻨﻋ و هرﺎﻤﺷ
ﻪﺤﻔﺻ
ﺶﻫوﮋﭘ تﺎﯿﻠﮐ :مود ﻞﺼﻓ
ﻞﮑﺷ -2 ﺑ يروﺮﻣ :1 ﻒﺸﮐ يﺎﻫﺪﮐرﺎﺑ ﯽﻧﺎﻣز ﺮﯿﺳ ﺮ هﺪﺷ
...
...
...
...
40
ﻞﮑﺷ -2 ﻦﯿﺑ ﻖﻃﺎﻨﻣ :2 ﻪﺘﺴﻫ ،ﯽﻧژ
يا ITS
. ...
. ...
...
...
51
ﺞﯾﺎﺘﻧ :مرﺎﻬﭼ ﻞﺼﻓ
ﻞﮑﺷ -4 ﺖﺧرد :1 يﺪﻧوﺎﺸﯾﻮﺧ ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﯽﺘﺳﻼﭘوﺮﻠﮐ ﯽﻟاﻮﺗ ﯽﻔﯾدر ﺎﻫﺎﯿﮔ ياﺮﺑrbcL
ن ﻦﯾﺮﯿﺷ نﺎﯿﺑ
،(آ) ﻦﺳﻮﺳ
غاﺮﭽﻠﭼ ﺑ ،(ب) (ه) ﺮﯿﺸﮐﺎﺧ ،(د) ﺐﻠﻌﺛ ،(ج) ﻪﻧﻮﺑﺎ
. ...
...
. ...
...
...
...
...
..
65
ﻞﮑﺷ -4 يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد :2 ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﯽﺘﺳﻼﭘوﺮﻠﮐ ﯽﻟاﻮﺗ ﯽﻔﯾدر rbcL
. ...
...
...
67
ﻞﮑﺷ -4 :3 يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﯽﺘﺳﻼﭘوﺮﻠﮐ ﯽﻟاﻮﺗ ﯽﻔﯾدر trnH-psbA
ﻪﻧﻮﺑﺎﺑ هﺎﯿﮔ ياﺮﺑ لﺎﻤﻋا نوﺪﺑ ،
Boot strapping . ...
...
...
...
70
ﻞﮑﺷ -4 :4 يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﯽﺘﺳﻼﭘوﺮﻠﮐ ﯽﻟاﻮﺗ ﯽﻔﯾدر trnH-psbA
و (آ) ﮓﻨﻫرﺎﺑ هﺎﯿﮔ ياﺮﺑ ﺮﯿﺸﮐﺎﺧ
(ب) . ...
...
...
..
70 ﻞﮑﺷ -4 :5 يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﯽﺘﺳﻼﭘوﺮﻠﮐ ﯽﻟاﻮﺗ ﯽﻔﯾدر trnH-psbA
. ...
...
...
71
ﻞﮑﺷ -4 :6 يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﯽﺘﺳﻼﭘوﺮﻠﮐ ﯽﻟاﻮﺗ ﯽﻔﯾدر هﺎﯿﮔ ياﺮﺑmatK
ﻪﻨﻣرد
. ...
. ...
....
. ...
73
ﻞﮑﺷ -4 :7 يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﻪﺘﺴﻫ ﯽﻟاﻮﺗ ﯽﻔﯾدر يا
ﻦﯾﺮﯿﺷ هﺎﯿﮔ ياﺮﺑITS (ب) ﻪﻧﻮﺑﺎﺑ ،(آ) نﺎﯿﺑ
ﺮﯿﺸﮐﺎﺧ و
ح
) ج (
. ...
...
...
...
...
...
...
74
ﻞﮑﺷ -4 :8 يﺪﻧوﺎﺸﯾﻮﺧ ﺖﺧرد ﻢﻫ ﺮﺑ ﯽﻨﺘﺒﻣNJ
ﻪﺘﺴﻫ ﯽﻟاﻮﺗ ﯽﻔﯾدر يا
ITS
...
. ...
...
...
75
ط
10
11
1
لوا ﻞﺼﻓ :
ﻪﻣﺪﻘﻣ
2
-1 ﻪﻣﺪﻘﻣ1 ﯽﮑﯾ ﯽﯾوراد نﺎﻫﺎﯿﮔ زا
ﻊﺑﺎﻨﻣ رﺎﯿﺴﺑ ﺪﻨﻤﺷزرا ﻊﯿﺳو هﺮﺘﺴﮔ رد ﻌﯿﺒﻃ
ﺖ ،ﺖﺸﮐ ،ﯽﻤﻠﻋ ﺖﺧﺎﻨﺷ ترﻮﺻ رد ﻪﮐ ﺪﻨﺘﺴﻫ
هﺮﻬﺑ و ﻪﻌﺳﻮﺗ ﯽﻣ ﺢﯿﺤﺻ يرادﺮﺑ
ﻮﺗ لﺎﻐﺘﺷا ،ﻪﻌﻣﺎﺟ ﺖﻣﻼﺳ رد ﯽﻤﻬﻣ ﺶﻘﻧ ﺪﻨﻧا ﺮﯿﻏ تاردﺎﺻ و ﯽﯾاز
ﺪﻨﺷﺎﺑ ﻪﺘﺷاد ﯽﺘﻔﻧ
،نارﺎﮑﻤﻫ و ﻦﮐﺪﯿﻔﺳ) (.1387
ظﺎﺤﻟ ﻪﺑ ﻢﻫ ﻊﻣاﻮﺟ ﯽﺘﻣﻼﺳ و ﺖﺷاﺪﻬﺑ ﻦﯿﻣﺄﺗ رد ﯽﺻﺎﺧ ﺖﯿﻤﻫا و شزرا زا ﯽﯾوراد نﺎﻫﺎﯿﮔ
يرﺎﻤﯿﺑ زا يﺮﯿﮕﺸﯿﭘ ﻢﻫ و نﺎﻣرد رادرﻮﺧﺮﺑ ﺎﻫ
.ﺪﻨﺘﺴﻫ ﻪﺘﺒﻟا هژاو نﺎﻫﺎﯿﮔ ﯽﯾوراد ﺎﻬﻨﺗ ﻪﺑ ﻦﯿﮑﺴﺗ هﺪﻨﻫد مﻻآ مدﺮﻣ قﻼﻃا
ﯽﻤﻧ دﻮﺷ ) ﺪﯿﻣا
،ﯽﮕﯿﺑ (1377 ﻪﮑﻠﺑ ﻦﯾا نﺎﻫﺎﯿﮔ رد ﺮﯾز هوﺮﮔ اﺬﻏ ﻪﺑ ناﻮﻨﻋ ﻢﻌﻃ هﺪﻨﻫد
،ﺎﻫ ﯽﻧﺪﯿﺷﻮﻧ
،ﺎﻫ ﻦﯾﺮﯿﺷ هﺪﻨﻨﮐ
،ﺎﻫ ﮓﻧر
ﯽﻌﯿﺒﻃ و هﺮﺸﺣ ﺶﮐ ﺎﻫ ) يرﻻﺎﺳ و
،نارﺎﮑﻤﻫ (1390
و ﻦﯿﻨﭽﻤﻫ ﻪﺑ ناﻮﻨﻋ هدﺎﻣ ﻪﯿﻟوا تﻻﻮﺼﺤﻣ ﯽﺸﯾارآ
و ﯽﺘﺷاﺪﻬﺑ ) يﺮﺒﮐا
،
(1392 ﯿﻧ ﺰ درﻮﻣ ا هدﺎﻔﺘﺳ راﺮﻗ ﯽﻣ ﺪﻧﺮﯿﮔ .
ﻤﻫا نﺎﻫﺎﯿﮔ ،ﯽﺨﯾرﺎﺗ ﺮﻈﻧ ﻪﻄﻘﻧ زا ﻪﺘﺷاد ﻊﻣاﻮﺟ ﻪﻌﺳﻮﺗ رد ﯽﻧاواﺮﻓ ﺖﯿ
ﺗ و ﺪﻧا هدروآﺮﻓ ﻦﺘﻓﺎﯾ ياﺮﺑ ﯽﻌﯿﺳو تﺎﻘﯿﻘﺤ و ﺎﻫ
ﺦﯾرﺎﺗ لﻮﻃ رد ﯽﻫﺎﯿﮔ ﯽﯾوراد ﯽﻌﯿﺒﻃ داﻮﻣ مﺎﺠﻧا
هﺪﺷ
، ﺮﺘﻤﮐ ﺎﻬﻨﺗ ﻪﮐ ﺖﺳﺎﺠﻨﯾا ﺖﯿﻤﻫا ﺰﺋﺎﺣ ﻪﺘﮑﻧ ﺎﻣا زا
ﺪﺻرد10 عﻮﻤﺠﻣ زا
250000 ﻧﻮﮔ
ﯽﻫﺎﯿﮔ ﻪ ﺖﺴﯾز دﺮﮑﻠﻤﻋ ﮏﯾ زا ﺶﯿﺑ ياﺮﺑ نﺎﻬﺟ درﻮﻣ و ﯽﯾﺎﺳﺎﻨﺷ ،ﯽﺘﺧﺎﻨﺷ
هدﺎﻔﺘﺳا ﻪﺘﻓﺮﮔ راﺮﻗ ﻪﺑ .ﺪﻧا
ترﺎﺒﻋ
ﺮﺑ ﺮﮕﯾد ﺮﺸﺘﻨﻣ يﺎﻫرﺎﻣآ سﺎﺳا هﺪﺷ
ﻂﺳﻮﺗ ﺖﺷاﺪﻬﺑ ﯽﻧﺎﻬﺟ نﺎﻣزﺎﺳ
، 1
ﻦﯿﺑ ﺎﻬﻨﺗ ﺎﺗ 35
لﻮﻃ رد ﯽﯾوراد هﺎﯿﮔ ﻪﻧﻮﮔ راﺰﻫ 70
ﺎﯾ ﮏﯾ ﻞﻗاﺪﺣ ياﺮﺑ نﺎﻣز ﺪﻨﭼ
درﻮﻣ فﺮﺼﻣ ﺑﻪ ﻪﺘﻓر رﺎﮐ ﺪﻧا . ﺒﻟا
،ﻪﺘ هدروآﺮﻓ فﺮﺼﻣ زا ﯽﻧﺎﻬﺟ ﺖﺷاﺪﻬﺑ نﺎﻣزﺎﺳ ﺖﯾﺎﻤﺣ يﺎﻫ
ﻪﺑ ،ﯽﻌﯿﺒﻃ ﻞﺑﺎﻗ ﻖﻧور زا لﻮﺼﺤﻣ ﻦﯾا ترﺎﺠﺗ و ﺪﯿﻟﻮﺗ ﻪﮐ ﺖﺳا هﺪﺷ ﺚﻋﺎﺑ دﻮﺧ ﻪﺑﻮﻧ
.ﺪﺷﺎﺑ رادرﻮﺧﺮﺑ نﺎﻬﺟ رد ﯽﻬﺟﻮﺗ
يروآدﻮﺳ ﻪﻧﺎﻔﺳﺄﺘﻣ يراﻮﮔﺎﻧ ﻞﺋﺎﺴﻣ و تﻼﮑﺸﻣ ،ﯽﯾوراد نﺎﻫﺎﯿﮔ ﯽﻧﺎﻬﺟ ترﺎﺠﺗ ﻪﺑ نوﺰﻓازور ﻪﺟﻮﺗ و يدﺎﺼﺘﻗا نﻼﮐ يﺎﻫ
ﺑ ﻊﺑﺎﻨﻣ ﻦﯾا ياﺮﺑ ار ﻪ
دﻮﺟو و هدروآ ﻪﻧﻮﮔ زا ﺎﯾ ضاﺮﻘﻧا ﺮﻄﺧ ﺎﺑ ار ﯽﻫﺎﯿﮔ يﺎﻫ ﺖﺳد
ﭗﯿﺗﻮﻧژ نداد ﻪﺘﺧﺎﺳ ﻪﺟاﻮﻣ ﯽﻣﻮﺑ يﺎﻫ
ﺖﺳا
،بﺎﻨﺑ ﯽﻔﺸﮐ) (1389
اﺮﯾز. ﺨﺑ ﻪﻧﻮﮔ ﻪﺑ طﻮﺑﺮﻣ ،ترﺎﺠﺗ زا ﯽﻤﯿﻈﻋ ﺶ ﻊﻤﺟ ﺖﻌﯿﺒﻃ زا ﻪﮐ ﺖﺳا ﯽﯾوراد ﯽﻫﺎﯿﮔ يﺎﻫ
يروآ هﺪﺷ
ﯽﺧﺮﺑ و هﻮﯿﺷ ﺎﺑ دﺎﻧ يﺎﻫ ﺑ ﺎﻬﻨﺗ ﻪﻧ ،ﺖﺳر ﻪﻧﻮﮔ ﻞﺴﻧ ضاﺮﻘﻧا ﻪ
ﯽﻣ ﺎﻫ ﻬﺟ و ﻪﻘﻄﻨﻣ ﯽﺘﺴﯾز عﻮﻨﺗ ﻪﮑﻠﺑ ﺪﻣﺎﺠﻧا ﺮﻄﺧ ﺎﺑ ﺰﯿﻧ ار نﺎ
ﯽﻣ ﻪﺟاﻮﻣ يدﻮﺑﺎﻧ .دزﺎﺳ
ﻧآ زا هﺮﯿﺧذ رﻮﺸﮐ ﺮﻫ ﯽﺘﺴﯾز عﻮﻨﺗ ﻪﮐ ﯽﯾﺎﺠ ﺐﯿﺳآ لﺎﺣ ﻦﯿﻋ رد و ﺪﻨﻤﺷزرا يا
.ﺖﺳا ﺮﯾﺬﭘ ﯽﯾﺎﺳﺎﻨﺷ
، ﻊﻤﺟ يروآ و
ﻪﻧﻮﻤﻧ ﻪﻌﻟﺎﻄﻣ مﺎﮔ ﻦﯿﺘﺴﺨﻧ ﻪﻠﻤﺟ زا ﯽﺘﺴﯾز يﺎﻫ
و ﺖﻇﺎﻔﺣ يﺎﺘﺳار رد ﺎﻫ هﺮﻬﺑ
ﯽﻤﻠﻋ يرادﺮﺑ زا ﯽﻫﺎﮔآ .ﺖﺳا ﯽﺘﺴﯾز عﻮﻨﺗ زا
World Health Organization
1
3
ﺸﻧاد ،نآ ﯽﯾﺎﺳﺎﻨﺷ ﯽﯾﺎﻧاﻮﺗ و ﯽﺘﺴﯾز عﻮﻨﺗ و ﺖﺳا لﻮﻤﺷ نﺎﻬﺟ ﯽ
رﻮﺸﮐ ﻪﯿﻠﮐ رد يروآدﻮﺳ ﺐﺟﻮﻣ ﯽﻣ نﺎﻬﺟ يﺎﻫ
.ددﺮﮔ و
اﺰﺑا ﯾر ﺖﺴ ﻪﺑ يرﻮﺸﮐ ﭻﯿﻫ ﻪﮐ ﯽﻤﻧ ﯽﯾﺎﻬﻨﺗ
ﺖﺳد نآ ﻪﺑ ﺪﻧاﻮﺗ ﺪﺑﺎﯾ
) و يرﻮﭽﺳا ﺮﺗاوﻮﺷ
،1
.(2011 عﻮﻨﺗ ﺎﺘﺳار ﻦﯿﻤﻫ رد ﻊﯿﺳو
ﯿﮔ نﻮﻨﻓ ﺪﻨﻣزﺎﯿﻧ ﯽﯾوراد نﺎﻫﺎ ور و
ش نآ ﯽﮑﯿﺘﻧژ عﻮﻨﺗ ﯽﺳرﺮﺑ و ﯽﯾﺎﺳﺎﻨﺷ ﺖﻬﺟ ﯽﺗوﺎﻔﺘﻣ يﺎﻫ ﯽﻣ ﺎﻫ
ﺪﺷﺎﺑ يرﺪﯿﺣ) ، .(1387
ﻪﻣﺎﻧﺮﺑ ﺖﯿﻟﺎﻌﻓ ﻪﻌﺳﻮﺗ ياﺮﺑ ﯽﻟﻮﺻا يﺰﯾر ﻖﯿﻗد ﯽﺳرﺮﺑ ﺪﻨﻣزﺎﯿﻧ ،ﯽﯾوراد نﺎﻫﺎﯿﮔ ﺶﺨﺑ رد ﺎﻫ
ﺖﺧﺎﻨﺷ ،دﻮﺟﻮﻣ ﺖﯿﻌﺿو
ﯽﻓﺎﮐ ﻊﺑﺎﻨﻣ ﻪﺻﺮﻋ رد دﻮﺟﻮﻣ ﻪﺑ ﯽﻌﯿﺒﻃ ﻊﺑﺎﻨﻣ و ﯽﻋارز يﺎﻫ
ﻪﻧﻮﮔ ﯽﻠﺻا هﺎﮕﺘﺳﺎﺧ ناﻮﻨﻋ ﺢﯿﺤﺻ ﺖﺧﺎﻨﺷ ﻦﯿﻨﭽﻤﻫ و ﯽﻣﻮﺑ يﺎﻫ
ﺖﯾدوﺪﺤﻣ زا ﺶﻟﺎﭼ و ﺎﻫ
ﯽﻣ ﺎﻫ ﺪﺷﺎﺑ ) ﻦﮐﺪﯿﻔﺳ و
،نارﺎﮑﻤﻫ ﻦﯾﺪﺑ .(1387
رد يﺮﯿﺑاﺪﺗ ﺪﯾﺎﺑ ﺐﯿﺗﺮﺗ ﺖﻬﺟ
و ﯽﯾﺎﺳﺎﻨﺷ ،ﻆﻔﺣ
ﺶﯿﭘ نﺎﻫﺎﯿﮔ ﻦﯾا ﺢﯿﺤﺻ فﺮﺼﻣ ﻪﻧﻮﮔ ضاﺮﻘﻧا ناﻮﺘﺑ ﺎﺗ دﻮﺷ ﯽﻨﯿﺑ
يﺎﻫ ﺎﺑ ﯽﯾوراد ا زر ش داد ﺶﻫﺎﮐ ار ﺎﯾ و
زا فﺮﺼﻣ ﻪﻧﻮﮔ يﺎﻫ
ياراد ﺖﺨﯾر ﯽﺳﺎﻨﺷ ﺑ ﻪﺑﺎﺸﻣ ﻪ ﯽﻣ ﺪﯾﺪﻬﺗ ار ﯽﻧﺎﺴﻧا ﯽﻨﻤﯾا ﺖﯿﻌﺿو ﻪﮐ ﻢﻫ يﺎﺟ
،ﺪﻨﻨﮐ دﻮﻤﻧ يﺮﯿﮔﻮﻠﺟ نارﺎﮑﻤﻫ و ﻦﭼ)
،2
.(2010 شور زا هزوﺮﻣا ﻪﻧﻮﮔ ﻖﯿﻗد ﯽﯾﺎﺳﺎﻨﺷ ياﺮﺑ ﯽﻔﻠﺘﺨﻣ يﺎﻫ
ﯽﻣ هدﺎﻔﺘﺳا ﯽﻫﺎﯿﮔ يﺎﻫ شور ﻦﯾا زا ﯽﮑﯾ .دﻮﺷ
ﯽﯾﺎﺳﺎﻨﺷ يﺎﻫ
ﮏﻤﮐ ﻪﺑ نﺎﻫﺎﯿﮔ ﯽﯾﺎﺳﺎﻨﺷ ،دﺎﻤﺘﻋا ﻞﺑﺎﻗ ﻓ
ﻦ ﺪﯾﺪﺟ يراﺬﮔﺪﮐرﺎﺑ
3DNA ﯽﻣ ﺪﺷﺎﺑ ﻪﮐ ﺖﺒﺴﻧ ﻪﺑ شور يﺎﻫ ﯽﺘﻨﺳ
،نازرا ﻊﯾﺮﺳ و
ياراد ﺖﻗد ﯽﯾﻻﺎﺑ رد ﺺﯿﺨﺸﺗ ﻪﻧﻮﮔ ﺎﻫ و ﻦﯿﻨﭽﻤﻫ رد ﯽﺑﺎﯾزرا عﻮﻨﺗ ﯽﺘﺴﯾز ﯽﻣ ﺷﺎﺑ ﺪ ،نارﺎﮑﻤﻫ و هداﺰﻧﺎﻣﺮﻬﻗ) (. 1391
ضﺮﻓ ﺮﺑ
ﻦﯾا ﺖﺳا ﻪﮐ ﯽﺧﺮﺑ نژ يﺎﻫ ﻪﺘﺴﻫ يا ﺎﯾ ﺪﯿﺘﺳﻼﭘ ي ﯽﻫﺎﯿﮔ ﺑ ﻪ ﻞﯿﻟد ﺖﻋﺮﺳ
ًﺎﺒﯾﺮﻘﺗ ﻦﯿﯾﺎﭘ ﻞﻣﺎﮑﺗ ردﺎﻗ ﻪﺑ ﮏﯿﮑﻔﺗ ﻪﻧﻮﮔ يﺎﻫ
ﯽﻫﺎﯿﮔ يراﺬﮔﺪﮐرﺎﺑ و ﺪﻨﺘﺴﻫ ﺑﻪDNA
ﻞﯿﻟد ﺰﯾﺎﻤﺗ ﻦﯿﺑ ﻪﻧﻮﮔ يا و مﺪﻋ ﺰﯾﺎﻤﺗ نورد ﻪﻧﻮﮔ يا راﺰﺑا ي يﻮﻗ رد ﯽﯾﺎﺳﺎﻨﺷ ﻪﻧﻮﮔ ﺎﻫ ﺑ ﻪ -
صﻮﺼﺧ ﻪﻧﻮﮔ يﺎﻫ ﯽﯾوراد ﯽﻣ ﺪﺷﺎﺑ ؛نارﺎﮑﻤﻫ و ﻦﭼ) (2010
.
ﻦﻓ ﻦﯾا لﺎﺳ زا ﺑ2003 ﻪ ﻪﻧﻮﮔ ﯽﯾﺎﺳﺎﻨﺷ رد ﺮﯿﮔاﺮﻓ رﻮﻃ ﻪﺑ ﯽﻫﺎﯿﮔ يﺎﻫ
ﻪﺘﻓﺮﮔ رﺎﮐ لﺎﺳ زا ﻪﮐ ﺪﺷ
مﻮﯿﺳﺮﺴﻨﮐ2004
) هﺪﻧز تادﻮﺟﻮﻣ ﺪﮐرﺎﺑ
4CBOL ( ﻪﺑ هار هﺪﻧز تادﻮﺟﻮﻣ ياﺮﺑ يراﺬﮔﺪﮐرﺎﺑ ﻪﻧﺎﺨﺑﺎﺘﮐ ﮏﯾ دﺎﺠﯾا رﻮﻈﻨﻣ .ﺪﺷ يزاﺪﻧا
يراﺬﮔﺪﮐرﺎﺑ هدﺎﻔﺘﺳا ﺎﺑDNA
زا ﯽﻟاﻮﺗ ﮏﯾ رﺎﯿﺴﺑDNA
دراﺪﻧﺎﺘﺳا هﺎﺗﻮﮐ ﻦﯿﺑ ﻪﻘﻄﻨﻣ ﺎﯾ نژ ﮏﯾ زا هﺪﺷ
ﯽﻧژ ﯽﻣ مﺎﺠﻧا دﺮﯿﮔ ﻪﮐ
ﮏﻤﮐ ﻪﺑ آ ن ﯽﻣ ﯽﻟاﻮﺗ ﯽﭘ ناﻮﺗ ﻪﻧﻮﮔ ﻪﭼ ﻪﺑ چرﺎﻗ ﺎﯾ هﺎﯿﮔ ،رﻮﻧﺎﺟ ﺮﻫ دﺮﺑ دراد ﻖﻠﻌﺗ يا
ﻨﯿﺘﻧاو) ﯽ نارﺎﮑﻤﻫ و ،5
(.2008 رد ﻪﺠﯿﺘﻧ
ﺎﺑ ﯽﻣ فﺪﻫ ﯽﻟاﻮﺗ ﺮﯿﺜﮑﺗ ﻪﻧﻮﮔ ناﻮﺗ
ﺑ ﮏﯾدﺰﻧ ﯽﻠﯿﺧ يﺎﻫ ﻪ
) داد ﺺﯿﺨﺸﺗ ﺮﮕﯾﺪﮑﯾ زا ار ﻢﻫ تﺮﺒﻫ
و نارﺎﮑﻤﻫ
، 6
(2003 . فﺪﻫ
ﻦﻓ ﻦﯾا ﯽﻠﺻا و ﻪﯿﻟوا ﺮﻫ ياﺮﺑ صﺎﺧ ﺪﮐرﺎﺑ ﯽﯾﺎﺳﺎﻨﺷ
ﯽﻣ هﺪﻧز تادﻮﺟﻮﻣ ﺪﮐرﺎﺑ ﯽﺗﺎﻋﻼﻃا ﮏﻧﺎﺑ ﻞﯿﮑﺸﺗ و ﻪﻧﻮﮔ مﺎﻤﺗ ﺎﺗ ﺪﺷﺎﺑ
هداد ﻦﯾا زا هدﺎﻔﺘﺳا و ﯽﺳﺮﺘﺳد نﺎﮑﻣا ﺎﯿﻧد ناﺮﮕﺸﻫوﮋﭘ هﺪﻧز تادﻮﺟﻮﻣ ﯽﺗﺎﻋﻼﻃا ﮏﻧﺎﺑ .ﺪﻨﺷﺎﺑ ﻪﺘﺷاد ار ﺎﻫ
7BOLD مﺎﻤﺗ
1 Showalter& Schori
2 Chen et al.
3 DNA barcoding
4 Consortium for the Barcode of Life
5 Valentini et al.
6 Hebert et al.
7 Barcode Of Life Data system
4
هﺎﯿﮔ ﻞﯿﺒﻗ زا مزﻻ تﺎﻋﻼﻃا ﻣ تﻻﺎﻘﻣ و ﺶﯾور ﻞﺤﻣ ،ﺪﺷر ﻂﯾاﺮﺷ ،ﯽﺳﺎﻨﺷ
هﺎﯿﮔ ﺎﺑ ﻂﺒﺗﺮ درﻮﻣ
ﯽﺳرﺮﺑ هداد يﺎﺟ دﻮﺧ رد ار
ﺖﺳا ﯽﺗﺎﻋﻼﻃا ﮏﻧﺎﺑ) (BOLD
.
ﯽﻧژ ﻪﻘﻄﻨﻣ تﺎﻧاﻮﯿﺣ ياﺮﺑ دﺎﻬﻨﺸﯿﭘ دراﺪﻧﺎﺘﺳا ﺪﮐرﺎﺑ ياﺮﺑ ﻪﮐ
ياراد هﺪﺷ نژ رد زﺎﺑ ﺖﻔﺟ658
ﺰﻣر هﺪﻨﻨﮐ موﺮﮐﻮﺘﯿﺳ
زاﺪﯿﺴﮐا )1 (coxI يرﺪﻨﮐﻮﺘﯿﻣ زا1
ﺖﺳا ﻪﻨﯿﻣز رد ﯽﻟو نﺎﻫﺎﯿﮔ
، ﺚﺤﺑ ﺖﯿﻌﺿو زا يرﺎﯿﺴﺑ و ﺖﺳا ﺰﯿﮕﻧاﺮﺑ
يﺎﻫدﺮﺒﻫار دﺎﻬﻨﺸﯿﭘ
-
ﺪﺷ ﺖﺳﻼﭘوﺮﻠﮐ زا دﺮﻔﻨﻣ ﻪﻘﻄﻨﻣ ﮏﯾ سﺎﺳاﺮﺑ ﺎﯾ ه نارﺎﮑﻤﻫ و سﺮﮐ)
،2
؛ 2005 نارﺎﮑﻤﻫ و يﺎﻫﻻ ،3
زا ﯽﺒﯿﮐﺮﺗ ﺎﯾ و (2008
ﯽﻣ ﻒﻠﺘﺨﻣ ﻖﻃﺎﻨﻣ ﺪﺷﺎﺑ
و سﺮﮐ) نﻮﺴﮑﯾرا ،4
؛ 2007 نارﺎﮑﻤﻫ و زﺎﭼ ،5
(2007 نﺎﮑﻣ ﻪﮐ ﻦﯿﺑ و ﯽﻧژ يﺎﻫ ﯽﺘﺳﻼﭘوﺮﻠﮐ ﯽﻧژ
ﻣ درﻮ تﺎﻌﻟﺎﻄﻣ رد هدﺎﻔﺘﺳا يﺪﻧوﺎﺸﯾﻮﺧ
ﻞﻣﺎﺷ نﺎﻫﺎﯿﮔ ﻖﻃﺎﻨﻣ
ﻧژ ﻞﯿﺒﻗ زا ﯽ ،matk
،rpoC ،rpoB ،accD ،ycFs ،Ndhy ﻪﻘﻄﻨﻣ
ﻪﺘﺴﻫ يا ﯽﻟاﻮﺗ وITS يﺎﻫ ﻦﯿﺑ ﯽﻧژ atp – F – atpH ،
tranl (uAA)-trnf (6AA) psbk – psbI ،
، trnH–psb ﯽﻣ
.ﺪﻨﺷﺎﺑ
ﻪﻨﯿﻣز رد ﯽﻔﻠﺘﺨﻣ نﺎﻘﻘﺤﻣ ،دﻮﺧ ﺶﻫوﮋﭘ
ﺑ ﻒﻠﺘﺨﻣ يﺎﻫﺪﮐرﺎﺑ زا هدﺎﻔﺘﺳا ﺎﺑ ﻪ
دﻮﺑ نﺎﻫﺎﯿﮔ رد ﻖﻓﻮﻣ ﯽﯾﺎﺳﺎﻨﺷ لﺎﺒﻧد ه
ﺪﻧا
) ،نارﺎﮑﻤﻫ و زﺎﭼ
؛2005 ﮓﻨﯿﺟ نارﺎﮑﻤﻫ و نﺎﯾ ،6
نارﺎﮑﻤﻫ و ﮓﻨﯿﻣ ؛2009 ،7
.(2011
ﻪﺑ ﯽﻣ ﺮﻈﻧ ﺎﺑ ﺪﺳر ﻦﯿﺑ و ﯽﻧژ ﻖﻃﺎﻨﻣ زا هدﺎﻔﺘﺳا ﯽﻧژ
ﯽﻣ نﺎﻫﺎﯿﮔ يراﺬﮔﺪﮐرﺎﺑ رد لواﺪﺘﻣ ياﺮﺑ ار ﯽﻟاﻮﺗ ﻦﯾﺮﺘﻬﺑ ناﻮﺗ
ﮔ يراﺬﮔﺪﮐرﺎﺑ ﺎﺑ ﻦﯿﻨﭽﻤﻫ .دﻮﻤﻧ ﯽﻓﺮﻌﻣ ﻞﯿﺑدرا نﺎﺘﺳا ﯽﯾوراد نﺎﻫﺎﯿﮔ يراﺬﮔﺪﮐرﺎﺑ ﻪﻧﻮ
هﺎﺒﺘﺷا ﯽﯾﺎﺳﺎﻨﺷ ﺮﻄﺧ ،ﯽﯾوراد يﺎﻫ
ﻪﻧﻮﮔ ﻪﻧﻮﮔ يراﺬﮔﺪﮐرﺎﺑ و ﺖﻓر ﺪﻫاﻮﺧ ﻦﯿﺑ زا ،ﻪﺑﺎﺸﻣ يﺎﻫ ﻪﻘﺒﻃ رد ار ﻖﻘﺤﻣ ،ﯽﻫﺎﯿﮔ يﺎﻫ
ﺢﯿﺤﺻ يﺪﻨﺑ ﻪﻧﻮﮔ
يﺎﻫ درﻮﻣ ﯽﺳرﺮﺑ
.داد ﺪﻫاﻮﺧ يرﺎﯾ ﻦﯾﺪﺑ رﻮﻈﻨﻣ ﺮﺿﺎﺣ ﻪﻌﻟﺎﻄﻣ رد ﯽﻫﺎﯿﮔ ﺞﯾار يﺎﻫﺪﮐرﺎﺑ ﯽﻟاﻮﺗ زا
ﻪﻠﻤﺟ زا
8ITS
9 ،
، rbcl
10matK
، trnH-psbA و
نﺎﻫﺎﯿﮔ
ﻪﻧﻮﭘ ،ﮓﻨﻫرﺎﺑ ﮐ ،ﻪﻧﻮﺑﺎﺑ ،ﺎﺳ
ﻦﺳﻮﺳ ،ﺐﻠﻌﺛ ،نارادﺎﻣﻮﺑ ،ﯽﻨﺳﺎ ﯿﺸﮐﺎﺧ ،ﻪﻨﻣرد ،غاﺮﭽﻠﭼ
ﻦﯾﺮﯿﺷ ،ﺮ نﺎﯿﺑ ا ﺪﯾدﺮﮔ هدﺎﻔﺘﺳ .
1 Cytochrome C Oxidase Subunit I
2 Kress et al.
3 Lahaye et al.
4 Kress & Erickson
5 Chase et al.
6 Jingyuan et al.
7 Ming et al.
8 Internal Transcribed Spacer
9 Ribulose Bisphosphate Carboxylase L
10 Maturase K
5
Family name: Fatemeh Name: Asadi
Title of Thesis :DNA barcoding of some Ardabil’s indigenous medicinal plants in order to documentation of their biodiversity.
Supervisors: Dr. Sara Dezhsetan and Dr. Robab Ghahremanzadeh Advisors: Dr. Jabraeil Razmjou and Dr. Mohammad Taghi Alebrahim Graduate Degree M.Sc.
Major: Agronomy and Plant Breeding Sciences Specialty: Plant Breeding University: Mohaghegh Ardabili Faculty: Agricultural Sciences Graduation date: 1394/3/27 Number of pages: 99 Abstract:
The correct use of medicinal plants requires the existence of detailed information. DNA barcoding technique is based on study and reconstruction phylogenetic relationship between organisms at different levels of taxonomy (species, genus, family, order, etc.) by using sequence data, however with increasing the number of studied genes, the more realistic understanding is obtained of phylogenetic relationship at different levels of taxonomy. This technique used to identify ten medicinal plants (Lilium ledebourii, Achillea millefolium, Glycyrrhiza glabra, Matricaria chamomilla, Artemisia spp., Plantago major, Cichorium intybus, Sisymbrium spp., Nepeta menthoides, Orchis spp.) collected from the Ardebil province. DNA extraction was done by the modified CTAB method and PCR was performed with primers designed based on Chloroplasts barcodes rbcL, trnH-psbA, matK and nuclear barcode, ITS. Then PCR products purified and sequenced. The percentage of prosperity amplification and sequencing barcodes were estimated in samples reviewed respectively, 70 and 50, 60 and 40, 50 and 10, 70 and 40. In order to evaluate ability level of each location barcode to separate studied plant species, several other sequences belong to the same genus were collected from NCBI database. The sequences with samples in NCBI database were blasting and Bioinformatics analysis were performed. In tree phylogeny, the species belonging to the same genus were separated based on rbcL and trnH-psbA barcode sequences. For barcode ITS, this was correct only in G. glabra. In this study, barcoding L. ledebourii with rbcL was done for first time. SNPs were counted for barcodes rbcL (less than 30), trnH-psbA (less than100) and matK (less than 20). But SNP of ITS barcode was only acceptable in G.
glabra and for other of samples of this barcode were counted more than 200. Genetic distance percentage were estimated low between studied plants sequences and sequences of species from same genus and barcode that collected from NCBI database for three barcodes of rbcL, matK and trnH-psbA, that is shown high similarity studied plants’ barcode sequence with sequences existed from same genus and barcode in the database. However, this percentage in ITS barcode was estimated low only for G. glabra sample. Thus, barcode rbcL due to high separation ability, the number of low SNP and universality in most species, was introduced as the best barcodes. However, trnH-psbA and ITS barcodes due to related problem with direct sequencing of PCR products and lack of access to high quality sequences, were identified as complementary barcodes. MatK barcode is not recommended for samples because of the low power amplification and sequencing.
Keywords: Medicinal plants, DNA Barcoding, Barcodes rbcL, trnH-psbA, matK and ITS
6
University of Mohaghegh Ardabili
Faculty of Agricultural Sciences
Department of Agronomy and Plant Breeding Sciences
Thesis submitted in partial fulfilment of the requirements for the degree of M.Sc. in Agricultural (Biotechnology)
Title:
DNA barcoding of some Ardabil’s indigenous medicinal plants in order to documentation of their biodiversity.
Supervisor(s):
Sara Dezhsetan (Ph. D)
Robab Ghahremanzadeh (Ph. D) Advisor(s):
Jabraeil Razmjou (Ph. D) Mohammad Taghi Alebrahim (Ph. D)
By:
Fatemeh Asadi June – 2015