Agrobacterium mediated Gene Transfer

UPDATED ON:- 01-07-2023

अप्रत्यक्ष जीन स्थानांतरण (Indirect Gene Transfer):- जब किसी जैविक कारक की सहायता से पादप कोशिकाओं के जीनोम में जीन को समावेशित करके स्थायी रूपान्तरण प्राप्त किया जाता है तो इसे अप्रत्यक्ष जीन स्थानांतरण कहते हैं। इसमें Agrobacterium जीवाणु का उपयोग किया जाता है।

(When stable transformation is achieved by incorporating genes into the genome of plant cells with the help of a biological factor, it is called indirect gene transfer. Agrobacterium bacterium is used in this.)

Agrobacterium मध्यस्थ जीन स्थानांतरण (Agrobacterium mediated Gene Transfer):-

•  Agrobacterium एक ग्राम –ve जीवाणु है जो पादप की जड़ों के पास स्थित मृदा में पाया जाता है।

(Agrobacterium is a gram -ve bacterium found in soil around plant roots.)

•  Agrobacterium जीवाणु की दो जातियाँ द्विबीजपत्री पादपों में रोग उत्पन्न करती हैं:-

(Two species of Agrobacterium bacteria cause diseases in dicot plants:-)

i. Agrobacterium tumefaciens

ii. Agrobacterium rhizogenes

i. Agrobacterium tumefaciens:- यह आवृतबीजी पौधों में Crown gall रोग उत्पन्न करता है। इसमें Ti – प्लाज़्मिड पाया जाता है।

(It causes Crown gall disease in angiosperm plants. Ti-plasmid is found in it.)

ii. Agrobacterium rhizogenes:- आवृतबीजी पौधों में Hairy root रोग उत्पन्न करता है। इसमें Ri – प्लाज़्मिड पाया जाता है।

(It cause hairy root disease in angiosperm plants. Ri - plasmid is found in it.)

•  Ti प्लाज़्मिड के T – DNA भाग में वांछित जीन को समावेशित कर पुनर्योजी DNA बनाया जाता है। जिसे Agrobacterium कोशिका में स्थानांतरित कर देते हैं।

(Recombinant DNA is made by incorporating the desired gene into the T - DNA portion of the Ti plasmid. Which are transferred to Agrobacterium cell.)

•  अब निर्जमित पर्ण के गोल टुकड़े काट लेते हैं।

(Now cut round pieces of sterilized leaf.)

•  अब इन टुकड़ों को Agrobacterium के साथ पूरी रात ऊष्मायन देते हैं।

(Now incubate these pieces with Agrobacterium overnight.)

•  अब 2 दिनों तक इन टुकड़ों को Shooting माध्यम में रखते हैं।

(Now kept these pieces in Shooting medium for 2 days.)

•  अब इस Shooting माध्यम में Kanamycin व Carbenicillin मिलाकर 3-4 सप्ताहों तक संवर्धित करते हैं।

(Now add Kanamycin and Carbenicillin in this shooting medium and cultured for 3-4 weeks.)

•  अब इसे Rooting माध्यम में स्थानांतरित करके Kanamycin व Carbenicilin डालकर 3-4 सप्ताहों तक संवर्धित करते हैं।

(Now transfer this to the rooting medium, add Kanamycin and Carbenicilin, culture it for 3-4 weeks.)

•  अब इस पौधे को गमले में Green house में स्थापित करते हैं।

(Now establish this plant in a pot in green house.)

•  कुछ दिनों पश्चात पौधे को खेत की मृदा में स्थापित कर देते हैं।

(After a few days, establish the plant in the soil of the field.)

जीन स्थानांतरण की विधि (Mechanism of Gene Transfer):-

•  Ti – प्लाज़्मिड के T – DNA भाग में यह गुण होता है कि यह पादप के जीनोम में समाकलित हो सकता है।

(The T - DNA portion of the Ti - plasmid has the property that it can integrate into the plant's genome.)

•  पर्ण के टुकड़ों से Aceto – Syringine रसायन निकलता है जो T – DNA के Vir – Operon को सक्रिय कर देता है।

(Leaf pieces release the Aceto - Syringine chemical that activates the Vir - Operon of T - DNA.)

•  सक्रिय होते ही T – DNA भाग प्लाज़्मिड से पृथक होकर बहुत सी पादप कोशिकाओं में प्रवेश कर जाता है।

(Once activated, the T-DNA portion separates from the plasmid and enters into many plant cells.)

•  अब यह T – DNA केंद्रक में जाकर जीनोम में समाकलित हो जाता है।

(Now this T-DNA integrates into the genome by going into the nucleus.)

•  इसके फलस्वरूप पादप का स्थायी रूपान्तरण हो जाता है।

(This results in stable transformation of the plant.)