heterologous expression of potato virus y coat protein, isolate pot187

نویسندگان

nemat sokhandan-bashir

mahin poorsmaile

mohammad hajizadeh

چکیده

background: the advent of recombinant dna technology has facilitated heterologous expression of proteins from various sources in different host systems including escherichia coli. if a plant virus coat protein is expressed in the bacterium it can be used as the antigen for antibody preparation. such a recombinant antigen preparation can be particularly useful where equipment such as ultracentrifuge is unavailable to purify virus particles to use as the antigen for conventional antibody preparation. objective: heterologous protein expression and purification of  the full length potato virus y (pvy) coat protein (cp) from isolate pot187 (an affiliate of strain n) to be used as an antigen was the aim of the study. materials and methods: reverse transcription polymerase chain reaction (rt-pcr) was carried out to amplify an 801 bp fragment of the cp gene from pvy-infected potato leaves. the amplicon was cloned into pgem-t easy. the cloned fragment was restricted by bamhi + saci and the purified fragment was cloned into the expression vector pet21a(+) which was restricted with the same enzymes. the generated plasmid was introduced into e. coli strain rosettatm. the expression was induced with isopropyl-b-d-thiogalactopyranoside (iptg) and its protein content was subjected to sds-page and western blotting. results: sds-page analysis of protein from the induced bacteria showed a ~35 kda protein corresponding to pvy cp. expression of the recombinant protein was confirmed by anti-his anitibody. conclusions: the full-length cdna of pvy-cp was amplified from the infected potato leaves. the cdna was heterologously expressed in e. coli. the produced recombinant cp can be used as an antigen to generate polyclonal antibody.

برای دانلود باید عضویت طلایی داشته باشید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Heterologous Expression of Potato Virus Y Coat Protein, Isolate Pot187

Background: The advent of recombinant DNA technology has facilitated heterologous expression of proteins from various sources in different host systems including Escherichia coli. If a plant virus coat protein is expressed in the bacterium it can be used as the antigen for antibody preparation. Such a recombinant antigen preparation can be particularly useful where equipment such as ultracentri...

متن کامل

Induction of Resistance to Potato Virus Y (PVY) Using Hairpin Construct of Coat Protein

Potato virus Y (PVY) is one of the most damaging viruses of potato plants which infecting most cultivars and causing significant yield and economical losses. The application of the concept of pathogen derived resistance opened new horizons for the development of virus-resistant plants. This research was carried out to study RNA silencing to engineered potato plants that are resistant to potato ...

متن کامل

A Novel Recombined Potato virus Y Isolate in China

This study reports the findings of a distinct Potato virus Y (PVY) isolate found in Northeast China. One hundred and ten samples (leaves and tubers) were collected from potato plants showing mosaic symptoms around the city of Harbin in Heilongjiang province of China. The collected tubers were planted and let to grow in a greenhouse. New potato plants generated from these tubers showed similar s...

متن کامل

Analysis of Potato virus Y Coat Protein Epitopes Recognized by Three Commercial Monoclonal Antibodies

BACKGROUND Potato virus Y (PVY, genus Potyvirus) causes substantial economic losses in solanaceous plants. Routine screening for PVY is an essential part of seed potato certification, and serological assays are often used. The commercial, commonly used monoclonal antibodies, MAb1128, MAb1129, and MAb1130, recognize the viral coat protein (CP) of PVY and distinguish PVYN strains from PVYO and PV...

متن کامل

Evidence for diversifying selection in Potato virus Y and in the coat protein of other potyviruses.

The modes of evolution of the proteins of Potato virus Y were investigated with a maximum-likelihood method based on estimation of the ratio between non-synonymous and synonymous substitution rates. Evidence for diversifying selection was obtained for the 6K2 protein (one amino acid position) and coat protein (24 amino acid positions). Amino acid sites in the coat proteins of other potyviruses ...

متن کامل

Expression of Prunus Necrotic Ringspot Virus Coat Protein in E. coli

Background and Amis: Serological assay is considered as one of the best choices for conducting large number of infection tests. Recombinant DNA technology has been used for expression of virus coat protein (CP) gene in prokaryotic bacterial cells such as Escherichia coli and the recombinant CP (rCP) is used as immunogen in antibody production. Heterologous CP protein expression and purification...

متن کامل

منابع من

با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید


عنوان ژورنال:
iranian journal of biotechnology

ناشر: national institute of genetic engineering and biotechnology

ISSN 1728-3043

دوره 13

شماره 4 2015

میزبانی شده توسط پلتفرم ابری doprax.com

copyright © 2015-2023