PRODUCTION OF PLANT ANTIBODIES AS A PLANT PROTECTION

PRODUCT
Lecture : Dr. Ir. Rukmowati Brotodjojo, M. Agr.

Arranged by :
Group 4
134210057 Hesti Ruhaimah
134210058 Widya S. A. Pangaribuan
134210059 Sisilia Maharani
134210060 Firdaus
134210064 Regina Valeria Barus

PA-A Class

AGROTECHNOLOGY STUDY PROGRAM

FACULTY OF AGRICULTURE
UPN “VETERAN” YOGYAKARTA
2023/2024
 CHAPTER I
INTRODUCTION

A. Background
Plant diseases are disorders or abnormalities that occur in plants
caused by various factors. Many factors that cause plant diseases are
caused by pathogens. Pathogens are living organisms, the majority of
which are microorganisms and are capable of causing disease in plants or
plants. Microorganisms include fungi, bacteria, viruses, mycoplasma
nematodes. There have been many efforts to control plant pest
organizations, one of which is to produce plant antibodies as plant
protection products using biotechnology.
Around 1990, plants were first considered as a potential host for
producing antibodies and the word “plantibody” was coined. The term
“plantibodies” describes the products of plants that have been genetically
engineered to express antibodies and antibody fragments. With this
technology, plants are being used as antibody factories (bioreactors),
utilizing their endomembrane and secretory systems to produce large
amounts of clinically viable proteins which can later be purified from the
plant tissue. Antibodies can be expressed in plants as either full-length
molecules or as smaller fragments. In essence, a plantibody is an antibody
produced by genetically modified plants. Antibodies, originally derived
from animals, are produced in plants by transforming the latter with
animal antibody genes. Although plants do not naturally make antibodies,
plantibodies have been shown to function in the same way as normal
antibodies. This concept of using plants as heterologous expression
systems for recombinant antibodies (plantibodies) is now more than two
decades old.
B. Research Problems
Based on the background that has been described above, the
problems that can occur are how are antibodies formed in plants for plant
protection against pathogens through biotechnology?

C. Objectives
The objective of this paper is Knowing the formation of antibodies
in plants for plant protection against pathogens through biotechnology.
 CHAPTER II
DISCUSSION

A. Antibody Structure
Antibodies (abbreviated Ab), also known as immunoglobulins
(abbreviated Ig), are large, Y-shaped proteins used by the immune system
to identify and neutralize foreign objects such as pathogenic bacteria and
viruses. Antibodies recognize the pathogen's unique molecules, called
antigens. Each "Y" end of the antibody contains a paratope (analogous to a
lock) that is specific for one particular epitope (analogous to a key) on the
antigen, allowing these two structures to bond precisely. Using this
binding mechanism, antibodies can mark infected microorganisms or cells
for attack by other components of the immune system or can neutralize
them directly (e.g. by blocking parts of the virus that are important for
invasion).
Antibodies are glycoproteins that belong to the immunoglobulin
superfamily. The terms antibody and immunoglobulin are often used
interchangeably,[1] although the term 'antibody' is sometimes used for
both secreted and soluble forms, in other words, excluding B cell
receptors.[6] Antibodies play a role in the body's immunity as preventing
pathogens from entering or damaging cells by binding to them, stimulating
the destruction of pathogens by coating them by macrophages and other
cells; and trigger pathogen damage by stimulating other immune responses
such as the complement pathway.
In humans and most mammals, an antibody unit consists of four
polypeptide chains, namely two identical heavy chains and two identical
light chains; they are linked by disulfide bonds. Each chain is a series of
domains: somewhat similar sequences each consisting of about 110 amino
acids. This domain is usually represented in a simplified schematic as a
rectangle. The light chain consists of one VL variable domain and one CL
 constant domain, while the heavy chain contains one VH variable domain
and three to four constant domains CH1, CH2, and so on.
Structurally, antibodies are also divided into two antigen-binding
fragments (Fab), each of which contains one VL, VH, CL, and CH1
domain, and a crystallizable fragment (Fc), which forms part of the stem
of the letter Y.[11] Between them there is a hinge region of the heavy
chain whose flexibility allows antibodies to bind epitope pairs at various
distances, to form molecular complexes (dimers, trimers, etc.), as well as
to bind effector molecules more easily.

B. Generating Pathogen - Specific Antibodies

Antibody-based resistance is a novel strategy for generating transgenic
plants resistant to pathogens. Decades ago it was shown that polyclonal
and monoclonal antibodies can neutralize viruses, bacteria and selected
fungi. This approach has been improved recently by the development of
recombinant antibodies (rAbs). There are some technology for generating
pathogens on plants with antibody spesific such as hybridoma and phage
display.
1. Hybridoma Technology
Hybridoma technology is the traditional way to generate
valuable monoclonal antibodies, and this involves the
immortalization of antigen-specific B-lymphocytes. The first
antibodies against plant pathogens were generated using hybridoma
technology, and have been expressed either as full-size
 immunoglobulins or as antibody fragments in plants. Hybridoma
technology allows the production of monoclonal antibodies (mAbs)
that are specific to individual genera, species or even isolates of
fungi. mAbs have been used successfully to detect, quantify and
visualize the saprotrophic growth of pathogens, such as
Rhizoctonia solani, in artificially and naturally infested soils and
have been used to quantify the effects of the hyperparasite
Trichoderma harzianum on the saprotrophic growth dynamics of
Rhizoctonia solani in compost-based systems.
The most appropriate antigens for the detection of fungi in
soils and composts were those that were extracellular and that were
constitutively expressed or could be induced. Studies of
extracellular antigen (enzyme) production in Trichoderma spp.
have shown that the enzyme β-1,3-glucanase presents an ideal
candidate for the production of mAbs specific to this genus. The
enzyme is extracellular and its production is constitutive or semi-
constitutive (Bull & Chesters, 1966; Elad et al., 1982; Ramot et al.,
2000, as cited in Thornton et al., 2002). In this study by Thornton
et al., 2002, they used a commercial β-1,3-glucanase preparation to
develop a murine hybridoma cell line secreting mAbs specific for
Trichoderma spp. and for fungi closely related to them. Using the
mAb MF2, they show how the antigen can be used as a molecular
marker for the detection and recovery of Trichoderma spp. in
naturally infested composts. This mAb could be used in
conjunction with nucleic-acid-based diagnostic techniques to aid in
the rapid detection, specific monitoring, recovery and identification
of Trichoderma spp.

2. Phage Display
Phage display was developed to mimic the key features of
antibody generation by the immune systems for isolating antibody
specificities. Recombinant DNA and molecular display
technologies have provided new opportunities for the creation of
recombinant antibodies. Phage display, involves the introduction of
peptide sequences (such as the antigen-binding domains of
recombinant antibodies) into the coat protein gene of a
bacteriophage so that multiple copies of the peptide are displayed
on the virion surface. Large peptide libraries, including diverse
libraries of antibodies, can be prepared using phage display and
screened by a process known as panning. Phage display can be
achieved using particles containing a whole phage genome plus the
inserted peptide. The phage particles may be used to transduce a
phagemid (a plasmid carrying the phage origin of replication and
one gene encoding a coat protein fusion). In the latter approach,
superinfection with a helper phage is necessary to package the
phagemid. This is preferable when the displayed protein is large
enough to reduce the infectivity of the phage particles, to avoid
packaging constraints. The minor coat protein pIII can tolerate N-
terminal fusions with scFvs, but larger peptides such as Fab
fragments are displayed more efficiently by fusion with the pVIII
major coat protein.
Phage display has been used by various plant virologists in
identification of peptides that bind to a pathogenic virus’s coat
protein. The phage display isolated peptides were very specific and
highly sensitive. At the very least, these have diagnostic potential
as they can be produced as fusions with proteins that serve as an
antigen for antibody-reporter molecule conjugates. They may also
constitute the basis for a novel, introduced disease resistance
strategy. Peptides with high affinity and specificity for vital viral
proteins could be identified, and subsequently, the capacity to
synthesize these peptides may be introduced into plants.
 In planta peptide production might prevent viral
proliferation in infected cells. Such a strategy has been used
successfully with antibodies, but antibody folding usually requires
an oxidizing environment conducive to forming specific
intracellular disulfide bonds necessary for function. Phage display-
selected peptides may not be so exacting in their requirements.
Indeed, phage display selected peptides capable of binding to a
coat protein of the rice black streaked dwarf virus (RBSDV), when
produced recombinantly for diagnostic purposes, have been shown
to also disrupt proper coat protein folding and reduce the
pathogenicity of RBSDV. Phage display has also assisted in the
elucidation of various host systems secunded to the virus to permit
successful infection and replication. Using the viral replication
enhancer protein, AC3 as bait, a phage library of random
dodecapeptides fused to a coat protein was panned to identify
interacting peptides that were then analyzed for homology to
proteins from the model plant, Arabidopsis thaliana. The revelation
of the pathways to which these proteins are integral has allowed a
more sophisticated understanding of events required for successful
viral lifecycle and the role of the multifunctional protein AC3 in
events leading to virus-induced gene silencing (Kushwaha et al.,
2012).
Numerous cloning and mutagenesis strategies have been
developed to create. preserve, and exploit maximal antibody
diversity. As in the immune system, isolated scFv genes can be
subjected further to mutation, and the mutants with more desirable
characteristics then can be selected. Phage display antibodies can
be used in the same range of applications as their hybridoma
counterparts, and moreover, can offer several advantages over
hybridoma methods. Phage antibody generation is rapid and can be
prepared in a few weeks using microgram levels of antigen. Phage
 display can produce antibodies that are not readily prepared by
hybridoma technology. These scFv antibodies can be manipulated
genetically with ease for the construction and expression of new
recombinant proteins, such as AFP-scFv fusions described below.
In addition, phage display hasbeen used also for the selection of
other proteins with binding properties (Simon et al., 2004).

Phage display cycle. 1) fusion proteins for a viral coat

protein + the gene to be evolved (typically an antibody fragment)
are expressed in bacteriophage. 2) the library of phage are washed
over an immobilised target. 3) the remaining high-affinity binders
are used to infect bacteria. 4) the genes encoding the high-affinity
binders are isolated. 5) those genes may have random mutations
introduced and used to perform another round of evolution. The
selection and amplification steps can be performed multiple times
at greater stringency to isolate higher-affinity binders.

C. Antibody - Based Resistance Against Plant Viruses

Plant viruses are biotrophic parasites, that is, they are not
considered living organisms outside the environment of plant cells. The
structure of a virus consists of nucleic acid and protein, viruses can infect
plants so that the plants become diseased. Symptoms of plants affected by
the virus can be seen including oak leaf patterns, chlorosis, and necrosis.
Viruses can be treated with antibodies, so that plants are resistant to
viruses.
Efforts to control viruses in plants can be done using one of the
biotechnological methods, namely genetic engineering, for example,
making transgenic plants. Antibody-based resistance is a new strategy to
produce transgenic plants that are resistant to plant viruses. Antibodies are
part of the immune system that work to protect the body from the dangers
of viruses, bacteria, germs, substances that can cause infectious diseases.
Making plant antibody products based on resistance to plant viruses can be
done in 2 ways, namely RNA silencing and genome editing.
1. Engineering RNA silencing-based resistance to viruses
RNA silencing strategies have been used in antiviral breeding
for more than thirty years, and many plants engineered to stably
transmit small RNAs targeting various viruses have been approved
for commercial release. RNA silencing is a sequence-specific RNA
degradation mechanism that occurs in a variety of eukaryotic
organisms including fungi (quelling), animals (RNA interference),
and plants (post-transcriptional gene silencing). RNA silencing,
also referred to as RNA interference (RNAi), is activated by the
presence of double-stranded RNA (dsRNA) molecules and induces
inhibition or suppression of gene expression in a nucleotide
sequence-specific manner (Hannon, 2002; Voinnet, 2005). In
plants, several major protein families are involved in RNA
silencing, including Dicer-like (DCL), Argonautes (AGO), RNA-
dependent RNA Polymerase (RDR) and Suppressor of Gene
Silencing (SGS). As type III RNases, DCL proteins process
dsRNA or miRNA precursors into siRNA or miRNA, respectively,
20 to 24-nt in length with two overhanging bases at the 3′ end.
These siRNAs or miRNAs are inserted into the endonuclease AGO
protein to form the RNA-induced silencing complex (RISC).
Directed by the containing siRNA/miRNA, RISC can bind to target
mRNA or noncoding RNA and then silence target gene expression
by cleaving the target RNA and carry out degradation, or recruit
DNA and histone modifiers and inhibit target gene transcription.
The cleaved target RNA can be recognized by RDR proteins that
amplify dsRNA to enhance the silencing effect. SGS proteins
stabilize the dsRNA substrate for DCL to produce secondary
siRNA and enhance the RNA silencing process

1. CRISPR/Cas (genome editing) based resistance engineering

against viruses.
Genome editing is a method that allows scientists to
change the DNA of many organisms, including plants, bacteria
and animals. Genome editing techniques not only integrate,
delete and/or mutate genes of interest, but also provide new
weapons against plant viruses. The CRISPR/Cas genome
editing system consists of a Cas protein endonuclease and a
single guide RNA (sgRNA) that directs the Cas protein to a
DNA or RNA target. Additionally, the sgRNA contains a
scaffold for Cas protein binding and a user-defined 20-nt long
spacer sequence for genome targeting. When CRISPR/Cas was
first explored, it succeeded in producing tobacco that was
resistant to Gemini virus.
 The mechanism of CISPR/CAS is transgenic or transit
expression of viruses targeting sgRNAs and their cognate Cas
proteins can effectively inhibit viral infection. Once DNA
viruses enter plant cells, for example geminiviruses, the viral
genome is converted into a double-stranded DNA
intermediate, which can be targeted and cleaved by the Cas9
protein of Streptococcus pyogenes (SpCas9). For RNA viruses,
Cas9 from Francisella novicida (FnCas9) and Cas13a have
been shown to confer effective viral resistance. Guided by
their cognate sgRNA or crRNA, FnCas9 and Cas13a can bind
to or cleave viral genomes or transcripts, respectively. The
viral genome is converted into a double-stranded DNA
intermediate, which can be targeted and cleaved by the Cas9
protein of Streptococcus pyogenes (SpCas9).

D. Antibody - Based Resistance Against Plant Bactery Pathogens

An example of gram-positive bacteria is the bacterium Clavibacter
michiganensis which causes ring rot disease in potatoes. Meanwhile,
gram-negative bacteria have more complex cell walls with less
peptidoglycan. An example of gram-negative bacteria is the bacteria
Xanthomonas oryzae which causes crackle disease in rice. Bacteria are
classified based on oxygen requirements into aerobic bacteria and
anaerobic bacteria. Aerobic bacteria are bacteria that need free oxygen to
get energy. An example of aerobic bacteria is the Ralstonia solanacearum
bacteria which causes wilt in tomato plants. Meanwhile, anaerobic bacteria
do not need free oxygen to get energy. An example of anaerobic bacteria is
the Pectobacterium carotovorum bacteria which causes wet rot in cabbage
plants. The classification of bacteria based on how they obtain food
(organic material) is divided into autotrophic and heterotrophic bacteria.
Autotrophic bacteria are bacteria that make their own food from inorganic
materials. Autotrophic bacteria, based on their energy source, are divided
into: photoautotrophs (energy sources from light) and chemoautotrophs
(energy sources from chemical reactions). Meanwhile, heterotrophic
bacteria are bacteria that do not prepare their own food but instead use
finished organic material that comes from other organisms. An example is
saprophytic bacteria which obtain food by decomposing the remains of
organisms.
Bacterial reproduction is divided into two, namely asexual and
sexual. Asexual reproduction of bacteria occurs through binary fission
where one bacterial cell divides into two daughter cells. The binary fission
process begins with the process of DNA replication into two identical
DNA copies, followed by cytoplasmic division and finally a dividing wall
is formed between the two bacterial daughter cells. Sexual reproduction in
bacteria is carried out in three ways, namely transformation, transduction
and conjugation. Transformation is sexual reproduction by transferring
DNA from one bacteria to another directly without a link. Transduction of
sexual reproduction by transferring DNA assisted by a phage virus as an
intermediary. If a bacterium is infected by a phage virus, the bacteria will
undergo lysis and release the phage and its DNA. The phage virus and
DNA then attach to other bacteria.
Conjugation is a stage of sexual reproduction in bacteria which is
characterized by the direct transfer of genetic material. The transfer occurs
from one bacteria to another via a conjugation bridge. Symptoms of a plant
being infected by bacteria are finding a rotten, slimy plant, it is possible
that this plant is being attacked by bacteria. In ornamental plants, it is
usually found at the base of rotting stems or withered plants. To prevent
infection, you should pay attention to plant care, don't water excessively,
especially if there is standing water. And don't overuse fertilizer.
The discovery of recombinants for controlling bacterial infections
by “Le Gall et al (1998) generated scFv fragments to recognize the major
immunodominant membrane protein (IMP) from the stolbur phytoplasma,
a mollicute that is largely restricted to the phloem elements of infected
 hosts. scFv is derived from the full-length monoclonal 2A10, which was
initially isolated from a hibrodome and expressed in E. coli whose
function is to ensure specification and stability of the scFv. ScFv-2A10
was then expressed constitutively in the cytosol of transgenic tobacco
plants using the CaMV 35S promoter, which is active in the phloem
among many other tissues. Research was carried out using transgenic
scions grafted onto infected tobacco rootstocks which remained
asymptomatic and flowered after 2 months, while control scions from wild
plants were infected by rootstocks and died before flowering.

E. Antibody-Based Resistence Against Fungal Plant Pathogens

Fungal plants is eukaryotic organisms with cell walls made of
chitin and do not have chlorophyll so that in obtaining food (nutrients)
fungi act as parasites (either obligate parasites or facultative parasites) or
as saprophytes (live by breaking down organic waste such as bankai into
inorganic material). Fungal structure of hyphae in fungi has several types,
namely as follows:
1. Hyphae that do not have a partition (aseptate hyphae), namely hyphae
that do not have a partition so that between one nucleus and another
there is no partition/membrane. Such hyphae are called senocytic.
2. Single-nucleated septate hyphae (uninuclear septate hyphae), namely
hyphae with cells that have a single nucleus. The septum divides the
hyphae into chambers with each chamber having one nucleus.
3. Septate hyphae have many nuclei (multinucleated septate hyphae),
namely hyphae with cells with many nuclei. The septum divides the
hyphae into chambers with multinucleated cells.
Antibody-Based Resistence Against Fungal Plant Pathogens found
by Peschen et al (2004) to prevent fungal diseases. They expressed a
fusion protein consisting of a recombinant scFv that recognizes a surface
protein present on Fusarium oxysporum f. sp. matthiolae combines with
antifungal proteins from Aspergillus giganteus. ScFv was isolated from a
phage display library derived from chickens immunized with fungal
spores. Preliminary analysis confirmed that the scFv-AFP fusion bound to
the fungus in vitro and inhibited its growth. Research on fungal pathogens
and their interactions with plants has identified several genes that confer
resistance to fungal pathogens. Pathogenic fungi encode antimicrobial
peptides, enzymes that synthesize antifungal metabolites, growth
inhibitors, proteins that inhibit fungal virulence, and proteins that induce
natural plant defenses including sensitive responses.
 CASE STUDY OF OPT MANAGEMENT BIOTECHNOLOGY
"Purification And Characterization Of A Viral Chitinase Active Against
Plant Pathogens And Herbivores From Transgenic Tobacco"

Previously, we demonstrated that transgenic tobacco plants expressing the

chitinase A from the Autographa californica nucleopolyhedrovirus virus
(AcMNPV ChiA) showed reduced damages after fungal pathogens and
lepidopteran larvae attack, without effects on a non-target insect population.
Those data indicated that the AcMNPV ChiA is an interesting candidate as a
molecule of biological origin for crop protection. Besides agricultural
applications, the enzymatic hydrolysis of chitin is a process of increasing interest
for the medical and industrial sectors such as the production of chito-
oligosaccharides and N-acetyl-d-glucosamine, the preparation of spheroplasts and
protoplasts from yeast and fungal species, and the bioconversion of chitin waste.
Furthermore chitinases also possess antibacterial, hypocholesterolemic and
antihypertensive activities and are also useful as food quality enhancers.
Tobacco transgenic lines expressing a chimeric ChiA gene were already
available. In these lines, the sequence coding the mature AcMNPV ChiA is fused,
at the 5' end, to a sequence coding a tobacco Signal Peptide, to target the
propeptide to the secretory pathway, and at the 3' end, to a sequence coding the
HDEL, as endoplasmic reticulum retention signal, and a myc epitope. The
transgene is under the control of the constitutive CaMV 35S RNA promoter.
Transgenic lines did not display any obvious phenotypic abnormalities and we
focused our attention on line 9, which accumulated the highest amount of
recombinant protein. An estimation of the amount of the rChiA protein produced
in transgenic plants was carried out by a fluorimetric immuno-assay calibrated on
a serial dilution of the PositopeTM, a recombinant protein engineered to contain,
among others the myc epitope. The constitutive expression of the recombinant
enzyme in transgenic tobacco plants was estimated to be about 14 mg kg−1 fresh
leaves weight (FLW, 0.2% of TSP). The chitinolytic activity of TSP from
transgenic and untransformed plants. In our assay conditions, the chitinolytic
activity of the transgenic extract increased linearly with the total protein
concentration. The isolation of the recombinant chitinase gave a yield of about 2
mg kg−1 FLW as assessed following chromatographic purification by SDS-PAGE
using known amounts of molecular weight markers as standards. The direct
confirmation of the identity of the isolated protein was obtained by sequencing its
N-terminus that corresponds to the amino acid sequence of the N-terminal region
of the mature AcMNPV ChiA (Ile-Pro-Gly-Thr-Pro-Val-Ile-Asp-Trp-Ala). The
sequence data proved also that, as expected, the Signal Peptide was correctly
removed.
Chitinases are hydrolytic enzymes produced by a vast range of organisms,
including insects, plants and animals. Because of their specific activity towards
chitin, they are considered to be highly selective, being for instance non-toxic to
higher vertebrates. For this reason, these enzymes have been long deemed
promising candidates as biopesticides and as enhancers of plant protection. In
addition, chitinases are also induced in plants in response to biotic stress and, as
implied by different studies, their heterologous expression should not have
detrimental effects on plant growth and development. The recombinant chitinase
was purified from tobacco leaves in which the enzyme was present at a
concentration of about 0.2% of TSP, a value that falls into the 0.1 - 1% of TSP
range typically observed for the proteins produced in nuclear transformants of
tobacco plants. This species was chosen as a non-food and non-feed crop that can
supply one of the most abundant leaf biomass per acre. The production of the
recombinant enzyme in tobacco plants overcomes some of the limitations of the
enzyme production in bacteria. Purified enzyme was 2 mg kg−1 FLW,
corresponding to a yield of 14%. It is likely that alternatives downstream
processing, which may include the use of a specific extraction buffer with
protease inhibitors and an affinity tag chromatography, could achieve higher
recovery. We showed that the purified recombinant enzyme displays a number of
interesting features. The rChiA, compared to the commercial enzyme, retains its
activity at higher pH values (>9), confirming its suitability as biopesticide against
herbivorous insects. The midgut environment of Lepidoptera larvae is usually at
pH values higher than nine, explaining why, among the chitinases so far tested,
only the insect moulting chitinase from Manduca sexta or the chitinase from
AcMNPV significatively perturbed larval growth of lepidopteran species. The
biological activity of the rChiA on herbivorous insects was experimentally
confirmed by its ability to increase the permeability of the PM of both B. mori and
H. virescens larvae. In addition, the enzyme showed antifungal activity towards A.
Alternata, a property that is shared by a number of microbial and plant chitinases.
However, in comparison to the S. marcescens chitinase A, the rChiA was more
effective in inhibiting germ tube elongation at lower doses. More interestingly,
while the optimum temperature of the enzyme was not different from that of
several chitinases, the rChiA outperformed the control enzyme in the stability
assay at 50◦C. This feature strongly increases rChiA value for industrial
processing of chitin, because current limitations on the use of chitinases are their
instability and their activity within a narrow temperature and pH range. In
conclusion, data indicated that the rChiA, purified with a reduced number of steps
from transgenic tobacco plants, could be very useful for the chitin industry, as
biopesticide, and for other biotechnological applications.
 CHAPTER III
CONCLUSION

1. Plant diseases are disorders or abnormalities that occur in plants caused by

various factors. Many factors that cause plant diseases are caused by
pathogens. Pathogens are living organisms, the majority of which are
microorganisms and are capable of causing disease in plants or plants.
Microorganisms include fungi, bacteria, viruses, mycoplasma nematodes.
There have been many efforts to control plant pest organizations, one of
which is to produce plant antibodies as plant protection products using
biotechnology.
2. Plant viruses are biotrophic parasites, that is, they are not considered living
organisms outside the environment of plant cells. The structure of a virus
consists of nucleic acid and protein, viruses can infect plants so that the
plants become diseased. Symptoms of plants affected by the virus can be
seen including oak leaf patterns, chlorosis, and necrosis. Viruses can be
treated with antibodies, so that plants are resistant to viruses.
3. The discovery of recombinants for controlling bacterial infections by “Le
Gall et al (1998) generated scFv fragments to recognize the major
immunodominant membrane protein (IMP) from the stolbur phytoplasma,
a mollicute that is largely restricted to the phloem elements of infected
hosts. scFv is derived from the full-length monoclonal 2A10, which was
initially isolated from a hibrodome and expressed in E. coli whose
function is to ensure specification and stability of the scFv. ScFv-2A10
was then expressed constitutively in the cytosol of transgenic tobacco
plants using the CaMV 35S promoter, which is active in the phloem
among many other tissues. Research was carried out using transgenic
scions grafted onto infected tobacco rootstocks which remained
asymptomatic and flowered after 2 months, while control scions from wild
plants were infected by rootstocks and died before flowering.
4. Antibody-based resistance is a novel strategy for generating transgenic
plants resistant to pathogens. Decades ago it was shown that polyclonal
and monoclonal antibodies can neutralize viruses, bacteria and selected
fungi. This approach has been improved recently by the development of
recombinant antibodies (rAbs). There are some technology for generating
pathogens on plants with antibody spesific such as hybridoma and phage
display.
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Zhao, Y., X. Yang, G. Zhou, T. Zhang. 2019. Engineering plant virus resistance:
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