Developing a real time PCR assay to screen and quantify HVA1 copy number in transgenic Moroccan durum wheat

مقدم أطروحة جامعية

Mukhtari, Wafa

مشرف أطروحة جامعية

al-Asli, Abd al-Ghani
al-Mustafa, al-Fahimi

أعضاء اللجنة

Sindidi, Khalid Sadaqa
Barradah, Fuad
Ibrahimi, Izz al-Din

الجامعة

جامعة الأخوين

الكلية

كلية الهندسة و العلوم

القسم الأكاديمي

التكنولوجيا الحيوية

دولة الجامعة

المغرب

الدرجة العلمية

ماجستير

تاريخ الدرجة العلمية

2010

الملخص الإنجليزي

Pasta Wheat (Triticum durum) is one of the leading commercial cereals worldwide and in Morocco because of the potential of its high yield.

In Morocco, drought is a real trauma that limits the growth and the yield of several crops including durum.

The Moroccan durum is considered land races species that must be maintained and valorized for the imporvement and sustainable cultivation.

Genetic transformation has become a widespread biotech-tool for making drought resistant transgenic for the sustainable increase in various crops production like durum wheat.

HVA1 barley gene has been one of the candidate genes to construct transgenic drought tolerant durum wheat.

Transgenic plants must be characterized at the molecular level because of the new DNA constituent.

At UATRS-CNRST (Centre National de Recherche Scientifique et Technique) Genomics platform the aim of our project was to analyze five transformed Triticum durum generation at the molecular level.

These transformants are obtained by introducing the foreign HVA1 gene of the Barley (Hordeum vulgare) into durum using ballistic-mediated transformation.

This genetic transformation is developed by researchers from plant biotechnology unit of the INRA (Rabat, Morocco).

Prior to implement PCR detection analysis we design specific primers that target the amplification of the gene HVA1.

Classical PCR was used thereafter to identify the new transformants.

The first and fifth transformants durum wheat progeny (F1 and F5) were identified as HVA1 transgenic.

Classical PCR also validates the specificity of our primers amplifying a unique 712bp band.

Next step was to develop a Real-time quantitative PCR (Q-PCR) assay for the absolute quantification of the HVA1 copy number in durum transgenic F5 generation.

The real time software quantifies 1.69 copies; therefore, two copies is the estimated HVA1 copy number that F5 progeny contains.

To validate this quantifying data, Q-PCR parameters including thresholds, efficiency, and linear regression were optimized during many real time PCR assays.

Transgenic copy number estimation is one of the primarily focus to characterize transgenic because it monitors the effectiveness of the transformation process.

Copy number determination can be very useful for the selection and cultivation of low copies of the foreign gene in transgenic seeds.

Infract, the transgenic grains might integrate the open crop market for their ration claimed in the norm.

التخصصات الرئيسية

الأحياء
النبات

الموضوعات

• علم الوراثة
• المغرب
• القمح القاسي

عدد الصفحات

62

قائمة المحتويات

• APA
• MLA
• AMA

لغة النص

الإنجليزية

نوع البيانات

رسائل جامعية

رقم السجل

BIM-646864