Tissue culture (Micropropagation)

(‫زراﻋﺔ اﻷﻧﺴﺠﺔ )اﻟﺘﻜﺎﺛﺮ اﻟﺪﻗﻴﻖ‬

 ‫ﻣﻘﺪﻣﺔ‬
‫™ ‪Micropropagation‬‬
‫¾ﺗﻨﻤﻴﺔ اﻟﺨﻼﻳﺎ أو اﻻﻧﺴﺠﺔ أواﻷﻋﻀﺎء اﻟﻨﺒﺎﺗﻴﺔ ﻋﻠﻰ ﺑﻴﺌﺔ‬
‫ﻏﺪاﺋﻴﺔ ﻓﻰ ﻇﺮوف ﻣﻌﻘﻤﺔ‬

‫™ ‪Totipotency‬‬
‫¾آﻞ ﺧﻠﻴﺔ ﻧﺒﺎﺗﻴﺔ ﺣﻴﺔ ﻟﺪﻳﻬﺎ اﻟﻘﺪرة ﻋﻠﻰ ﺗﻜﻮﻳﻦ ﻧﺒﺎت‬
‫آﺎﻣﻞ‬
 ‫ﻣﻤﻴﺰات وﺗﻄﺒﻴﻘﺎت اﻟﺘﻜﺎﺛﺮ اﻟﺪﻗﻴﻖ‬

‫اﻻﻧﺘﺎج اﻟﻐﺰﻳﺮ ﻟﻠﻨﺒﺎﺗﺎت ‪Mass production‬‬ ‫¾‬

‫اﻧﺘﺎج ﻧﺒﺎﺗﺎت ﺧﺎﻟﻴﺔ ﻣﻦ اﻻﻣﺮاض واﻟﻔﻴﺮوﺳﺎت‬ ‫¾‬

‫اآﺜﺎر اﻟﻨﺒﺎﺗﺎت اﻟﺼﻌﺒﺔ اﻻآﺜﺎر‬ ‫¾‬

‫اﻧﺘﺎج اﺻﻨﺎف ﺟﺪﻳﺪة دات ﺻﻔﺎت ﺗﺴﻮﻳﻘﻴﺔ ﻣﻤﺘﺎزة‬ ‫¾‬

‫اﻟﻬﻨﺪﺳﺔ اﻟﻮراﺛﻴﺔ وﺗﻄﺒﻴﻘﺎﺗﻬﺎ ﻓﻰ ﻣﺠﺎل ﺗﺤﺴﻴﻦ اﻟﻨﺒﺎﺗﺎت‬ ‫¾‬

‫زراﻋﺔ اﻷﺟﻨﺔ اﻟﻐﻴﺮ ﻧﺎﺿﺠﺔ‬ ‫¾‬

‫اﻟﺘﻮﻓﻴﺮ ﻓﻰ اﻟﻮﻗﺖ اﻟﻼزم ﻟﻠﺘﺮﺑﻴﺔ ﺑﺎﻟﻄﺮق اﻟﺘﻘﻠﻴﺪﻳﺔ‬ ‫¾‬

‫اﻟﺘﻐﻠﺐ ﻋﻠﻰ ﻣﺸﺎآﻞ اﻟﺴﻜﻮن وﻋﺪم اﻟﺘﻮاﻓﻖ‬ ‫¾‬

‫اﻧﺘﺎج ﻧﺒﺎﺗﺎت اﺣﺎدﻳﺔ وﺑﺎﻟﺘﺎﻟﻰ ﻧﺒﺎﺗﺎت اﺻﻴﻠﺔ وراﺛﻴﺎ‬ ‫¾‬

‫ﻋﻴﻮب وﻣﺤﺪدات اﻟﺘﻜﺎﺛﺮ اﻟﺪﻗﻴﻖ‬

Expense ‫ﻣﻜﻠﻒ‬ ƒ

Species specifity ‫ﺧﺼﻮﺻﻴﺔ اﻻﻧﻮاع‬ ƒ

Contamination ‫اﻟﺘﻠﻮث‬ ƒ

Variability ‫اﻻﺧﺘﻼف‬ ƒ

Acclimatation ‫اﻻﻗﻠﻤﺔ‬ ƒ
 ‫اﻟﺘﻘﻨﻴﺎت اﻟﻤﺴﺘﺨﺪﻣﺔ ﻻآﺜﺎر اﻟﻨﺒﺎﺗﺎت‬
structure Regeneration Explant source

Plantlet Axillary shoot Meristem or shoot tip

Adventitious shoot Leaf pieces or stem

intraneod
seedling Seed cultures Seeds

Embryo cultures Mature or immature

embryo
callus Callus cultures Vegetative tissue

Protoplast cultures Single cells

Somatic embryo Direct or indirect Embryo, seedling or

leaf
 ‫ﻣﻜﻮﻧﺎت ﻣﻌﻤﻞ زراﻋﺔ اﻻﻧﺴﺠﺔ‬

Culture room

Laminar flow Autoclave

hood

Media preparation room

‫ﻣﻜﻮﻧﺎت اﻟﺒﻴﺌﺎت اﻟﻐﺪاﺋﻴﺔ‬

Inorganic salts ‫™ أﻣﻼح ﻣﻌﺪﻧﻴﺔ‬

Organic compounds ‫™ ﻣﺮآﺒﺎت ﻋﻀﻮﻳﺔ‬
Sucrose™
Vitamins™
Growth regulators™
Supports ‫™ ﻣﻮاد ﻣﺪﻋﻤﺔ‬
Agar ‫™اﺟﺎر‬
 ‫ﻣﻨﻈﻤﺎت اﻟﻨﻤﻮ ‪Growth regulators‬‬
‫ﺗﻀﺎف ﺑﻜﻤﻴﺎت ﺻﻐﻴﺮة ﻟﻠﺒﻴﺌﺎت ﻟﺘﻨﺸﻴﻂ ﻧﻤﻮ اﻟﺴﻴﻘﺎن واﻟﺠﺪور وﻣﻨﻬﺎ‪:‬‬
‫‪ o‬اﻻآﺴﻴﻨﺎت ‪Auxins‬‬
‫‪IAA, IBA, NAA o‬‬
‫‪ o‬اﻟﺴﻴﺘﻮآﻴﻨﻴﻨﺎت ‪Cytokinins‬‬
‫‪BA, Kinetin, zeatin o‬‬
‫‪ o‬اﻟﺠﺒﺮﻳﻠﻴﻨﺎت ‪Gibberellins‬‬
‫‪Gibberellic acid o‬‬

‫ﺑﻌﺾ اﻻآﺴﻴﻨﺎت واﻟﺴﻴﺘﻮآﻴﻨﻴﻨﺎت ﻻ ﺗﺪوب ﻓﻰ اﻟﻤﺎء ﻟﺪا ﺗﺴﺘﺨﺪم ﻗﻄﺮات‬

‫ﻣﻦ ‪ NaOH‬ﻟﻼآﺴﻴﻨﺎت أو ‪ HCl‬ﻟﻠﺴﻴﺘﻮآﻴﻨﻴﻨﺎت وﻗﺪ ﻳﺴﺘﺨﺪم‬
‫اﻻﻳﺜﺎﻧﻮل ﻟﻜﻠﻴﻬﻤﺎ‬
‫اﻷدوات‬

‫اﻧﺎﺑﻴﺐ اﺧﺘﺒﺎر‬ ‫¾‬

‫أﻃﺒﺎق ﺑﺘﺮى‬ ‫¾‬

‫ﻋﻠﺐ زراﻋﺔ‬ ‫¾‬

‫زﺟﺎﺟﺎت‬ ‫¾‬

‫ﻣﺨﺎﺑﻴﺮ‬ ‫¾‬

‫ادوات زراﻋﺔ‬ ‫¾‬

‫ﻣﺎﺻﺎت‬ ‫¾‬

‫ﻣﻮازﻳﻦ‬ ‫¾‬

‫ﻣﻘﻴﺎس ‪Ph‬‬ ‫¾‬

‫ﻣﻘﻠﺐ وﺳﺨﺎن‬ ‫¾‬

‫اﻟﺘﺤﻜﻢ ﻓﻰ ﻇﺮو ف اﻟﺰراﻋﺔ‬

‫™ اﻟﺤﺮارة‬
‫™ ﺷﺪة اﻟﻀﻮء‬
‫™ ﻃﻮل اﻟﻔﺘﺮة اﻟﻀﻮﺋﻴﺔ‬
‫™ ﺟﻮدة اﻟﻀﻮء‬
‫™ اﻟﺘﻬﻮﻳﺔ‬
 ‫ﻣﺮاﺣﻞ اﻟﺘﻜﺎﺛﺮ اﻟﺪﻗﻴﻖ‬
Culture establishment ‫ﻣﺮﺣﻠﺔ اﻻﻧﺸﺎء‬ (1

Multiplication stage ‫ﻣﺮﺣﻠﺔ اﻟﺘﻀﺎﻋﻒ‬ (2

Root formation ‫ﻣﺮﺣﻠﺔ ﺗﻜﻮﻳﻦ اﻟﺠﺪور‬ (3

Acclimatization stage ‫ﻣﺮﺣﻠﺔ اﻻﻗﻠﻤﺔ‬ (4

‫ﻣﺮﺣﻠﺔ ﺑﺪء اﻟﺰراﻋﺔ ‪Establishment stage‬‬
‫اﻟﻐﺮض ﻣﻨﻬﺎ ﻓﺼﻞ اﻟﻨﺴﻴﺞ اﻟﻨﺒﺎﺗﻰ ‪ explant‬ووﺿﻌﻪ ﻋﻠﻰ‬ ‫™‬
‫اﻟﺒﻴﺌﺔ ﻻﻧﺘﺎج ﺳﻴﻘﺎن‬
‫ﺗﺸﺘﻤﻞ هﺪﻩ اﻟﻤﺮﺣﻠﺔ ﻋﻠﻰ‪:‬‬ ‫¾‬

‫اﺧﺘﻴﺎر اﻟﻨﺴﻴﺞ وﺗﻌﻘﻴﻤﻪ‬

‫اﻋﺪاد اﻟﺒﻴﺌﺔ وﺗﻌﻘﻴﻤﻬﺎ‬
‫وﺿﻊ اﻟﻨﺴﻴﺞ ﻋﻠﻰ اﻟﺒﻴﺌﺔ ﻓﻰ ﻇﺮوف ﻣﻌﻘﻤﺔ‬
‫ﺣﻔﻆ اﻟﺰراﻋﺎت ﻓﻰ ﻇﺮوف ﺛﺎﺑﺘﺔ‬
 ‫ﻣﺮﺣﻠﺔ اﻟﺘﻀﺎﻋﻒ ‪Multiplication stage‬‬

‫™ ﺗﻬﺪف اﻟﻰ ﻣﻀﺎﻋﻔﺔ أﻋﺪاد اﻟﺴﻴﻘﺎن اﻟﻨﺎﺗﺠﺔ ﻋﻠﻰ اﻟﺒﻴﺌﺔ‬

‫ﺑﺎﻟﻤﻌﻤﻞ ﻟﻠﻌﺪد اﻟﻤﺮاد ﺗﺠﺪﻳﺮﻩ‬
‫ﺗﺸﺘﻤﻞ هﺪﻩ اﻟﻤﺮﺣﻠﺔ ﻋﻠﻰ‪:‬‬ ‫¾‬

‫اﺳﺘﺨﺪام ﻣﻨﻈﻤﺎت ﻧﻤﻮ ﻟﻠﺘﻀﺎﻋﻒ‬

‫ﻧﻘﻞ اﻟﺰراﻋﺎت ﻟﺒﻴﺌﺎت ﻃﺎزﺟﺔ ‪Subcultures‬‬
‫ﻣﺮﺣﻠﺔ ﺗﻜﻮﻳﻦ اﻟﺠﺪور ‪Root formation stage‬‬

‫™ ﺗﻬﺪف اﻟﻰ دﻓﻊ اﻟﺴﻴﻘﺎن اﻟﻰ اﻟﺘﺠﺪﻳﺮ واﻋﺪادهﺎ ﻟﻤﺮﺣﻠﺔ‬

‫زراﻋﺘﻬﺎ ﺑﺎﻟﺨﺎرج ‪ex vitro‬‬
‫ﺗﺸﺘﻤﻞ هﺪﻩ اﻟﻤﺮﺣﻠﺔ ﻋﻠﻰ‪:‬‬ ‫¾‬

‫اﺳﺘﺨﺪام ﺑﻴﺌﺔ ﻣﻨﺎﺳﺒﺔ ﻟﻠﺘﺠﺪﻳﺮ‬

‫اﻟﺘﻬﻴﺌﺔ ﻟﻠﺰراﻋﺔ ‪ex vitro‬‬
 ‫ﻣﺮﺣﻠﺔ اﻻﻗﻠﻤﺔ ‪Acclimatization stage‬‬

‫™ اﻟﻐﺮض ﻣﻨﻬﺎ ﻧﻘﻞ اﻟﻨﺒﺎﺗﺎت اﻟﻤﺠﺪرة ﻣﻦ اﻟﺒﻴﺌﺔ اﻟﻐﺪاﺋﻴﺔ‬

‫اﻟﻤﻌﻘﻤﺔ ﺑﺎﻟﻤﻌﻤﻞ اﻟﻰ اﻟﺘﺮﺑﺔ اﻟﻌﺎدﻳﺔ ﺑﺎﻟﺨﺎرج ‪Ex vitro‬‬
‫ﺗﺸﺘﻤﻞ هﺪﻩ اﻟﻤﺮﺣﻠﺔ ﻋﻠﻰ‪:‬‬ ‫¾‬

‫ﻧﻘﻞ اﻟﻨﺒﺎﺗﺎت ﻣﻦ اﻟﺒﻴﺌﺔ اﻟﻰ اﻟﺘﺮﺑﺔ‬

‫اﻻﻗﻠﻤﺔ ﺑﺎﻟﺰراﻋﺔ ﺑﺘﺮﺑﺔ ﻣﻌﻴﻨﺔ ﺗﺤﺖ ﻇﺮوف ﻣﻌﻴﻨﺔ‬
‫‪Organogensis and somatic embryogensis‬‬

‫™ ‪Organogensis‬‬
‫¾ ﻋﻤﻠﻴﺔ ﺗﻜﻮﻳﻦ ﺳﻴﻘﺎن و‪/‬أو ﺟﺪور ﻣﻦ اﻟﺨﻼﻳﺎ أو اﻻﻧﺴﺠﺔ‬
‫ﻣﺒﺎﺷﺮ دون اﻟﻤﺮور ﺑﻤﺮﺣﻠﺔ اﻟﻜﺎﻟﻮس ‪callus‬‬ ‫ƒ‬

‫ﻏﻴﺮ ﻣﺒﺎﺷﺮ ﺑﺘﻜﻮﻳﻦ آﺎﻟﻮس ﻗﺒﻞ اﻟﻤﺘﻜﺸﻒ ﻟﺴﻴﻘﺎن‬ ‫ƒ‬

‫™ ُ‪Embryogensis‬‬
‫¾ ﻋﻤﻠﻴﺔ ﺗﻨﻤﻴﺔ أو اﻧﺘﺎج أﺟﻨﺔ ﻣﻦ ﺧﻼﻳﺎ ﺟﺴﺪﻳﺔ أو ﺧﻀﺮﻳﺔ‬
‫ﺑﺪﻻ آﻦ اﺗﺤﺎد اﻟﺠﺎﻣﻴﻄﺎت اﻟﻤﺪآﺮة واﻟﻤﺆﻧﺜﺔ‬
‫‪Biotechnology and Genetic engeneering‬‬
‫اﻟﻬﻨﺪﺳﺔ اﻟﻮراﺛﻴﺔ‬
‫ﻋﻤﻠﻴﺔ ﻧﻘﻞ اﻟﺼﻔﺎت )اﻟﺠﻴﻨﺎت( ﻣﻦ أى آﺎﺋﻦ ﺣﻰ اﻟﻰ آﺎﺋﻦ ﺣﻰ اﺧﺮ‬
‫ﺗﺤﻮﻳﺮ ﻣﺒﺎﺷﺮ‪ :‬ﻧﻘﻞ ﺍﻟﺠﻴﻨﺎﺕ ﻣﺒﺎﺷﺮﺓ ﻟﻠﺨﻼﻳﺎ ﺍﻟﻨﺒﺎﺗﻴﺔ‬
‫ﺗﺤﻮﻳﺮ ﻏﻴﺮ ﻣﺒﺎﺷﺮ‪ :‬ﺍﺳﺘﺨﺪﺍﻡ ﺍﻟﺒﻜﺘﻴﺮﻳﺎ ﻛﻨﺎﻗﻞ ﻟﻠﺠﻴﻨﺎﺕ‬
Protoplast fusion ‫دﻣﺞ اﻟﺒﺮوﺗﻮﺑﻼﺳﺖ‬