Detection of Erwinia spp. on potato (Solanum tuberosum l.)‎ using conventional and PCR technique

العناوين الأخرى

التحري عن بكتيريا Erwinia spp. على البطاطا (Solanum tuberosum L.)‎ باستخدام الطرق التقليدية و تقنية تفاعل إنزيم البلمرة التسلسلي

مقدم أطروحة جامعية

Muhammad, Maytham Jamil

مشرف أطروحة جامعية

Salman, Ihab Dawud

الجامعة

جامعة بغداد

الكلية

كلية العلوم

القسم الأكاديمي

قسم التقانة الإحيائية

دولة الجامعة

العراق

الدرجة العلمية

ماجستير

تاريخ الدرجة العلمية

2013

الملخص الإنجليزي

A total of seventy samples of infected potato tubers were collected from Baghdad city during the period from September 2011 to February 2012.

These samples were collected from Alradwaniyah potato’s store/ Albyaa market and from Awrad Alnahar potato’s store in Abu ghraib.

As a standard Erwinia strains ATCC 15713 and ATCC 495 were provided from Plant Protection department/ College of Agriculture/ Baghdad University.

The Crystal Violet Pectate (CVP) was used as a selective- diagnostic medium to isolate and purify Erwinia under the specific conditions necessary for the cultivation of these bacteria (temperature and growth period…etc.).

The strains and isolates of Erwinia were characterized according to the morphological and microscopic characteristics, along with the biochemical tests and API 20E tests; which indicates that the bacteria are gram negative, rod shaped, peritrichously flagellated and facultative anaerobic.

The colonies form characteristic deep cups-like cavities or pits.

The results determining the optimal temperature and pH for the growth of local Erwinia isolates have revealed that the temperature at 30 ºC and pH (7.4) were suitable to obtain profuse growth of Erwinia.

Results of biochemical tests showed that Erwinia isolates have a positive reaction for catalase, raffinose and cellobiose and a negative reaction for oxidase and sorbitol, which means that all isolates were within the two Erwinia species Erwinia carotovora and Erwinia chrysanthemi.

The Virulence determination tests of local Erwinia isolates were determined by growing the local Erwinia isolates on PDA medium, the best six local Erwinia isolates II were chosen according to their containing rate and their spreading on the surface of the media, then these local Erwinia isolates were tested using Potato tuber’s slices to show their efficiency to infect the potatoes.

Agarose gel electrophoresis was performed to detect plasmid profile for the selected isolates and standard strains of Erwinia by using salting out method for total DNA extraction.

The results showed that all the six selected Erwinia isolates and the two standard strains Erwinia carotovora subsp.

carotovora ATCC 15713 and ATCC 495 contained at least two small plasmid DNA bands, whereas the Miniprep method was not reproducible for plasmid DNA extraction, the results also revealed that all local isolates exhibited no bands of any plasmid.

By using the specific DNA primer (Ec001) for Erwinia Erwinia carotovora subsp.

carotovora, this primer has a sequence F 5′-CGGTTACGATCAGCGTCTCG-3′ and R 5′-GATGTGCCGATGCCGATAC-3′.

The local Erwinia isolates were determined by using primer (Ec001) and PCR technique.

Amplification products were tested with 1.0 % agarose gel and stained with ethidium bromide.

The results showed that the selected local isolates of Erwinia and Erwinia carotovora subsp.

carotovora ATCC 15713 potato’s pathogenic had a positive reaction (one band 312 bp) for the former primer (Ec001) and by depending on the biochemical tests and gel electrophoresis, results found that five of local Erwinia isolates resembles Erwinia carotovora subsp.

carotovora a pathogenic for potato, but the sixth Erwinia isolate and Erwinia carotovora subsp.

carotovora ATCC 495 potato’s nonpathogenic had a negative reaction (no band), this indicated that the isolate was nonpathogenic for potatoes or belongs to another Erwinia species.

التخصصات الرئيسية

الأحياء

عدد الصفحات

89

قائمة المحتويات

• APA
• MLA
• AMA

لغة النص

الإنجليزية

نوع البيانات

رسائل جامعية

رقم السجل

BIM-601173