Journal of Crop Protection
Journal of Crop Protection
JCP
Agriculture
http://jcp.modares.ac.ir
1
admin
2251-9041
2251-905X
en
jalali
1398
5
1
gregorian
2019
8
1
8
4
online
1
fulltext
en
Development and molecular analyses of Xanthomonas pthA specific scFv recombinant monoclonal antibodies
پژوهشی اصیل
Original Research
The <em>Xanthomonas</em> <em>citri</em> pv. <em>citri</em> (<em>Xcc</em>) is causal agent of bacterial citrus canker which is major disease of citrus throughout the world. The pthA bacterial effector protein is presented within the infected plants and indispensable of canker. The scFv antibodies are valuable tools for diagnosis and suppression of pathogens within plants. The present article describes developing and characterization of specific recombinant monoclonal scFv antibodies against pthA effector protein. For this aim, the gene encoding pthA protein was heterologously expressed in <em>Escherichia</em> <em>coli</em> and used for screening of Tomlinson phage display antibody library to pinpoint specific single chain variable fragment (scFv). In each round of panning, the affinity of phage towards pthA was checked by enzyme linked immunosorbent assay (ELISA). The data was indicative of about 50% of the monoclonal phages to be reactive strongly against pthA protein. Among the positive clones, 5 samples (A12, B8, C1, H8 and G8) were capable of detecting <em>Xcc</em>-infected plant samples and recombinant pthA protein. Restriction fragment length polymorphism showed similar banding pattern for all 5 scFvs as renamed to pthA-scFG8. HB2151 <em>E.</em> <em>coli</em> cells were infected by the phage bearing pthA-scFG8, and the expression of the peptide was induced by IPTG to produce a 30 kDa recombinant molecule. I-TASSER was used for homology modeling of both scFv and pthA and docking was carried out by Hex program. The latter demonstrated binding energy of −784 kcal/mol in scFv-pthA.
Biopanning, citrus bacterial canker, phage display, single chain fragment variable
417
429
http://jcp.modares.ac.ir/browse.php?a_code=A-10-13604-2&slc_lang=en&sid=3
Hamideh
Raeisi
ha.raeesi@gmail.com
1003194753284600136911
1003194753284600136911
No
Department of Plant Protection, Faculty of Agricultural Sciences, Guilan University, Rasht, Iran.
Mohammad Reza
Safarnejad
mrsafarnejad@yahoo.com
1003194753284600136912
1003194753284600136912
Yes
Department of Plant Viruses, Iranian Research Institute of Plant Protection, Agricultural Research Education and Extension Organization of Iran (AREEO), Tehran, Iran.
Seyed Mehdi
Alavi
mehdi.alavi@protonmail.com
1003194753284600136913
1003194753284600136913
No
Department of Plant Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.
Naser
Farrokhi
n_farrokhi@sbu.ac.ir
1003194753284600136914
1003194753284600136914
No
Department of Cell & Molecular Biology, Faculty of Life Sciences & Biotechnology, Shahid Beheshti University G. C, Tehran, Iran.
Seyed Ali
Elahinia
a_elahinia@yahoo.com
1003194753284600136915
1003194753284600136915
No
Department of Plant Protection, Faculty of Agricultural Sciences, Guilan University, Rasht, Iran.
Hossein
Safarpour
safarpour701@yahoo.com
1003194753284600136916
1003194753284600136916
No
Cellular and Molecular Research Center, Birjand University of Medical Sciences, Birjand, Iran.
Farshid
Sharifian
f.sharifian@ut.ac.ir
1003194753284600136917
1003194753284600136917
No
Department of Plant Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.