Polymerase chain reaction The polymerase chain reaction(PCR) is ascientific technique inmolecular

biology toamplify a single or a few copies of a piece ofDNA across several orders of magnitude, generating thousands to millions of copies of a particular DNA sequence. Developed in 1983 by Kary Mullis,[1] PCR is now a common and often indispensable technique used
[2][3]

in

medical

and

biological

research

labs

for

variety

of

applications.

These include DNA cloning for sequencing, DNA-based phylogeny, or

functional analysis of genes; the diagnosis ofhereditary diseases; the identification of genetic fingerprints (used inforensic sciences and paternity testing); and the detection and diagnosis of infectious diseases. In 1993, Mullis was awarded the Nobel Prize in Chemistry along with Michael Smith for his work on PCR.[4] The method relies on thermal cycling, consisting of cycles of repeated heating and cooling of the reaction for DNA melting and enzymaticreplication of the DNA. Primers (short DNA fragments) containing sequences complementary to the target region along with a DNA polymerase (after which the method is named) are key components to enable selective and repeated amplification. As PCR progresses, the DNA generated is itself used as a template for replication, setting in motion a chain reaction in which the DNA template is exponentiallyamplified. PCR can be extensively modified to perform a wide array ofgenetic manipulations. Almost all PCR applications employ a heat-stable DNA polymerase, such as Taq polymerase, an enzyme originally isolated from the bacteriumThermus aquaticus. This DNA polymerase enzymatically assembles a new DNA strand from DNA building-blocks, the nucleotides, by using single-stranded DNA as a template and DNA oligonucleotides (also calledDNA primers), which are required for initiation of DNA synthesis. The vast majority of PCR methods use thermal cycling, i.e., alternately heating and cooling the PCR sample to a defined series of temperature steps. These thermal cycling steps are necessary first to physically separate the two strands in a DNA double helix at a high temperature in a process called DNA melting. At a lower temperature, each strand is then used as the template in DNA synthesis by the DNA polymerase to selectively amplify the target DNA. The selectivity of PCR results from the use of primers that are complementary to the DNA region targeted for amplification under specific thermal cycling conditions. PCR principles and procedure PCR is used to amplify a specific region of a DNA strand (the DNA target). Most PCR methods typically amplify DNA fragments of up to ~10 kilo base pairs (kb), although some techniques allow for amplification of fragments up to 40 kb in size.[5] A basic PCR set up requires several components and reagents.[6] These components include:

DNA template that contains the DNA region (target) to be amplified. Two primers that are complementary to the 3'(three prime) ends of each of the sense and anti-sense strand of the DNA target. Taq polymerase or another DNA polymerase with a temperature optimum at around 70 C. Deoxynucleoside triphosphates (dNTPs; nucleotides containing triphosphate groups), the building-blocks from which the DNA polymerase synthesizes a new DNA strand. Buffer solution, providing a suitable chemical environment for optimum activity and stability of the DNA polymerase. Divalent cations, magnesium or manganese ions; generally Mg2+ is used, but Mn2+ can be utilized for PCR-mediated DNA mutagenesis, as higher Mn2+ concentration increases the error rate during DNA synthesis[7] Monovalent cation potassium ions.

The PCR is commonly carried out in a reaction volume of 10200 l in small reaction tubes (0.20.5 ml volumes) in a thermal cycler. The thermal cycler heats and cools the reaction tubes to achieve the temperatures required at each step of the reaction (see below). Many modern thermal cyclers make use of the Peltier effect, which permits both heating and cooling of the block holding the PCR tubes simply by reversing the electric current. Thinwalled reaction tubes permit favorable thermal conductivity to allow for rapid thermal equilibration. Most thermal cyclers have heated lids to prevent condensation at the top of the reaction tube. Older thermocyclers lacking a heated lid require a layer of oil on top of the reaction mixture or a ball of wax inside the tube. Procedure

Figure 2: Schematic drawing of the PCR cycle. (1) Denaturing at 9496 C. (2) Annealing at ~65 C (3) Elongation at 72 C. Four cycles are shown here. The blue lines represent the

DNA template to which primers (red arrows) anneal that are extended by the DNA polymerase (light green circles), to give shorter DNA products (green lines), which themselves are used as templates as PCR progresses. Typically, PCR consists of a series of 20-40 repeated temperature changes, called cycles, with each cycle commonly consisting of 2-3 discrete temperature steps, usually three (Fig. 2). The cycling is often preceded by a single temperature step (called hold) at a high temperature (>90C), and followed by one hold at the end for final product extension or brief storage. The temperatures used and the length of time they are applied in each cycle depend on a variety of parameters. These include the enzyme used for DNA synthesis, the concentration of divalent ions and dNTPs in the reaction, and the melting temperature (Tm) of the primers.[8]

Initialization step: This step consists of heating the reaction to a temperature of 9496 C (or 98 C if extremely thermostable polymerases are used), which is held for 19 minutes. It is only required for DNA polymerases that require heat activation by hot-start PCR.[9] Denaturation step: This step is the first regular cycling event and consists of heating the reaction to 9498 C for 2030 seconds. It causes DNA melting of the DNA template by disrupting the hydrogen bonds between complementary bases, yielding single-stranded DNA molecules. Annealing step: The reaction temperature is lowered to 5065 C for 2040 seconds allowing annealing of the primers to the single-stranded DNA template. Typically the annealing temperature is about 3-5 degrees Celsius below the Tm of the primers used. Stable DNA-DNA hydrogen bonds are only formed when the primer sequence very closely matches the template sequence. The polymerase binds to the primer-template hybrid and begins DNA formation . Extension/elongation step: The temperature at this step depends on the DNA polymerase used; Taq polymerase has its optimum activitytemperature at 75 80 C,[10][11] and commonly a temperature of 72 C is used with this enzyme. At this step the DNA polymerase synthesizes a new DNA strand complementary to the DNA template strand by adding dNTPs that are complementary to the template in 5' to 3' direction, condensing the 5'-phosphate group of the dNTPs with the 3'-hydroxyl group at the end of the nascent (extending) DNA strand. The extension time depends both on the DNA polymerase used and on the length of the DNA fragment to be amplified. As a ruleof-thumb, at its optimum temperature, the DNA polymerase will polymerize a thousand bases per minute. Under optimum conditions, i.e., if there are no limitations due to limiting substrates or reagents, at each extension step, the amount of DNA target is doubled, leading to exponential (geometric) amplification of the specific DNA fragment. Final elongation: This single step is occasionally performed at a temperature of 70 74 C for 515 minutes after the last PCR cycle to ensure that any remaining singlestranded DNA is fully extended. Final hold: This step at 415 C for an indefinite time may be employed for short-term storage of the reaction.

To check whether the PCR generated the anticipated DNA fragment (also sometimes referred to as the amplimer oramplicon),agarose gel electrophoresisis employed for size

separation of the PCR products. The size(s) of PCR products is determined by comparison with a DNA ladder (a molecular weight marker), which contains DNA fragments of known size, run on the gel alongside the PCR products .

ELECTROPHORETIC KARYOTYPING
Electrophoretic karyotyping is a term first introduced by Carle and Olson in 1985 ( 1) to describe the use of the new technique of pulsed field gel electrophoresis (PFGE) to visualize whole chromosomes from unicellular organisms. Conventional agarose gel electrophoresis has a useful upper size limit of about 20 kilobases (kb). PFGE extends this limit to at least 5.7 megabases (Mb) (2) and may be up to 12 Mb (3), encompassing the size range of chromosomes from many bacteria, fungi, and protozoa. Chromosomes from most of these organisms fail to condense sufficiently during mitosis to allow karotyping by light microscopy and, prior to the advent of PFGE, estimates of genome size and chromosome number were based on genetic linkage analysis, DNA reassociation kinetics, and in some cases, electron microscopy.

Elicitors
Are defined as substances of biotic origin which induce a defence response. Biochemically They were originally defined by Noel Keen as inducers of phytoalexin biosynthesis (people only knew about phytoalexins in those days). This definition now also covers inducers of:

hypersensitive response lignification PR proteins

Various substances are better considered to be signalling substances or hormones rather than elicitors e.g. salicylic acid, ethylene, jasmonates. Examples of elicitors

Proteins, peptides Carbohydrates: beta-glucans (especially heptaglucan), xylans oligogalacturans (especially 10 to 15-mers) chitosan (>hexamers) Fatty acids, glycosyl lipids

Abiotic elicitors It include UV light, heavy metals

Ex : The air pollutant ozone has recently been found to resemble fungal elicitors it can induce plant signal molecules such as ethylene and salicylic acid, as well as certain genes and biosynthetic pathways associated with pathogen and oxidative defence. The action of ambient ozone on the plant defence system may predispose the plant to enhanced attack by pathogens, but may also lead to induced resistance. The results mean that ozone can be regarded as a new experimental tool for analyzing stress responses.

Biotoc elicitors
Glycoprotein and Oligosaccharides These elicitor molecules are highly specific structures, that at very low concentrations, induce plant defense responses. Glycoproteins are proteins covalently attached to carbohydrates. Covalent means that a number of electron pairs of a protein atom are shared with electron pairs of a carbohydrate atom to form the specific glycoprotein. All of the Grigg Brothers "Green Spec" granular fertilizers contain specific glycoprotein's which elicit or stimulate the production of mycorrhizal and other fungi in soils, stimulate the plant immune system, and assist in stress recovery.

Mycorrhizal fungi act as plant root extensions, bringing in more water and nutrients. The increase in soil fungi increases the organic biomass which makes the soil less prone to compaction and opens up capillary spaces for air and water. These fungi are Mother Nature's form of aeration. More oxygen penetration will reduce anaerobic abiotoc stresses such as black layer. Oligosaccharides from fungal and plant cell walls have been scientifically shown to elicit plant defense responses. All of the Grigg Brothers "Green Spec" granular fertilizers contain specific oligosaccharides, Oligosaccharide elicitors generally have sugar residues. Chitin and chitosan oligosaccharide elicitors may act as potent elicitors. Little is known about the mechanisms responsible for percieving these oligosaccharide signals and transducing this information to the plant, so that defense genes can be turned on. The specific glycoproteins and oligosaccharides contained in the "Green Spec" fertilizers are: hepta-betaglucoside and chitin oligosaccharide.

Signal transduction occurs when an extracellular signaling molecule activates a cell

surface receptor. In turn, this receptor alters intracellular molecules creating a response. two stages in this process: 1. A signaling molecule activates a specific receptor on the cell membrane. 2. A second messenger transmits the signal into the cell, eliciting a physiological response. In either step, the signal can be amplified. Thus, one signalling molecule can cause many [2] responses.
[1]

There are

Extracellular
Extracellular receptors are integral transmembrane proteins and make up most receptors. They span the plasma membrane of the cell, with one part of the receptor on the outside of the cell and the other on the inside. Signal transduction occurs as a result of a ligand binding to the outside; the molecule does not pass through the membrane. This binding stimulates a series of events inside the cell; different types of receptor stimulate different responses and receptors typically respond to only the binding of a specific ligand. Upon binding, the ligand induces a change in theconformation of the [22] inside part of the receptor. These result in either the activation of an enzyme in the receptor or the exposure of a binding site for other intracellular signaling proteins within the cell, eventually propagating the signal through the cytoplasm. In eukaryotic cells, most intracellular proteins activated by a ligand/receptor interaction possess an enzymatic activity; examples include tyrosine kinase and phosphatases. Some of them create second messengers such as cyclic AMP and IP3, the latter controlling the release of intracellular calcium stores into the cytoplasm. Other activated proteins interact with adaptor proteins that facilitate signalling protein interactions and coordination of signalling complexes necessary to respond to a particular stimulus. Enzymes and adaptor proteins are both responsive to various second messenger molecules. Many adaptor proteins and enzymes activated as part of signal transduction possess specialized protein domains that bind to specific secondary messenger molecules. For example, calcium ions bind to the EF hand domains of calmodulin, allowing it to bind and activate calmodulindependent kinase. PIP3 and other phosphoinositides do the same thing to the Pleckstrin homology domains of proteins such as the kinase protein AKT.

[edit]G protein-coupled For more details on this topic, see G-protein-coupled receptor. G protein-coupled receptors (GPCRs) are a family of integral transmembrane proteins that possess seven transmembrane domains and are linked to a heterotrimeric G protein. Many receptors are in this family, including adrenergic receptors and chemokine receptors. Signal transduction by a GPCR begins with an inactive G protein coupled to the receptor; it exists as a heterotrimer consisting of G, G, and G. Once the GPCR recognizes a ligand, the conformation of the receptor changes to activate the G protein, causing G to bind a molecule of GTP and dissociate from the other two G-protein subunits. The dissociation exposes sites on the subunits that [23] can interact with other molecules. The activated G protein subunits detach from the receptor and initiate signalling from many downstream effector proteins such asphospholipases and ion channels, [24] the latter permitting the release of second messenger molecules. The total strength of signal amplification by a GPCR is determined by the lifetimes of the ligand-receptor complex and receptoreffector protein complex and the deactivation time of the activated receptor and effectors through intrinsic enzymatic activity. A study was conducted where a point mutation was inserted into the gene encoding the chemokine receptor CXCR2; mutated cells underwent a malignant transformation due to the expressionof CXCR2 in an active conformation despite the absence of chemokine-binding. This [25] meant that chemokine receptors participate in cancer development. [edit]Tyrosine and histidine kinase Receptor tyrosine kinases (RTKs) are transmembrane proteins with an intracellular kinase domain and an extracellular domain that binds ligands; examples include growth factor receptors such as [26] the insulin receptor. To perform signal transduction, RTKs need to form dimers in the plasma [27] membrane; the dimer is stabilized by ligands binding to the receptor. The interaction between the cytoplasmic domains stimulates the autophosphorylation of tyrosines within the domains of the RTKs, causing conformational changes. The receptors' kinase domains are subsequently activated, initiating phosphorylation signaling cascades of downstream cytoplasmic molecules that facilitate [26] various cellular processes such as cell differentiation andmetabolism. As is the case with GPCRs, proteins that bind GTP play a major role in signal transduction from the activated RTK into the cell. In this case, the G proteins are members of the Ras, Rho, and Raf families, referred to collectively as small G proteins. They act as molecular switches usually tethered to membranes by isoprenyl groups linked to their carboxyl ends. Upon activation, they assign proteins to specific membrane subdomains where they participate in signaling. Activated RTKs in turn activate small G proteins that activate guanine nucleotide exchange factors such asSOS1. Once activated, these exchange factors can activate more small G proteins, thus amplifying the receptor's initial signal. The mutation of certain RTK genes, as with that of GPCRs, can result in the expression of [28] receptors that exist in a constitutively-activate state; such mutated genes may act as oncogenes. Histidine-specific protein kinases are structurally distinct from other protein kinases and are found in prokaryotes, fungi, and plants as part of a two-component signal transduction mechanism: a phosphate group from ATP is first added to a histidine residue within the kinase, then transferred to an aspartate residue on a receiver domain on a different protein or the kinase itself, thus activating [29] the aspartate residue. [edit]Integrin For more details on this topic, see Integrin.

Integrins are produced by a wide variety of cells; they play a role in cell attachment to other cells and the extracellular matrix and in the transduction of signals from extracellular matrix components such as fibronectin and collagen. Ligand binding to the extracellular domain of integrins changes the protein's conformation, clustering it at the cell membrane to initiate signal transduction. Integrins lack kinase activity; hence integrin-mediated signal transduction is achieved through a variety of intracellular protein kinases and adaptor molecules, the main coordinator being integrin-linked [30] kinase. As shown in the picture to the right, cooperative integrin-RTK signalling determines the timing of cellular survival, apoptosis, proliferation, and differentiation. Important differences exist between integrin-signalling in circulating blood cells and non-circulating cells such as epithelial cells; integrins of circulating cells are normally inactive. For example, cell membrane integrins on circulating leukocytes are maintained in an inactive state to avoid epithelial cell attachment; they are only activated in response to stimuli such as those received at the site of aninflammatory response. In a similar manner, integrins at the cell membrane of circulating platelets are normally kept inactive to avoidthrombosis. Epithelial cells (which are noncirculating) normally have active integrins at their cell membrane, helping maintain their stable [31] adhesion to underlying stromal cells that provide signals to maintain normal functioning. [edit]Toll gate For more details on this topic, see toll-like receptor. When activated, toll-like receptors (TLRs) take adapter molecules within the cytoplasm of cells in order to propagate a signal. Four adaptor molecules are known to be involved in signaling, which [32][33][34] are Myd88, TIRAP, TRIF, and TRAM. These adapters activate other intracellular molecules such as IRAK1, IRAK4,TBK1, and IKKi that amplify the signal, eventually leading to the induction or suppression of genes that cause certain responses. Thousands of genes are activated by TLR signaling, implying that this method constitutes an important gateway for gene modulation. [edit]Ligand-gated ion channel For more details on this topic, see ligand-gated ion channel. A ligand-gated ion channel, upon binding with a ligand, changes conformation to open a channel in the cell membrane through which ions relaying signals can pass. An example of this mechanism is found in the receiving cell of a neural synapse. The influx of ions that occurs in response to these channels opening induces action potentials, such as those that travel along nerves, by depolarizing the membrane of post-synaptic cells, resulting in the opening of voltage-gated ion channels. An example of an ion allowed into the cell during a ligand-gated ion channel opening is Ca ; it acts as a second messenger initiating signal transduction cascades and altering the physiology of the responding cell. This results in amplification of the synapse response between synaptic cells by remodelling the dendritic spines involved in the synapse. [edit]Intracellular Further information: Intracellular receptor Intracellular receptors, such as nuclear receptors and cytoplasmic receptors, are soluble proteins localized within their respective areas. The typical ligands for nuclear receptors are lipophilichormones like the steroid hormones testosterone and progesterone and derivatives of vitamins A and D. To initiate signal transduction, the ligand must pass through the plasma membrane by passive diffusion. On binding with the receptor, the ligands pass through the nuclear membrane into the nucleus, enabling gene transcription and protein production.
2+

Activated nuclear receptors attach to the DNA at receptor-specific hormone-responsive element (HRE) sequences, located in the promoter region of the genes activated by the hormone-receptor complex. Due to them enabling gene transcription, they are alternatively called inductors of gene expression. Activation of gene transcription is slower than signals directly affecting existing proteins; therefore, the effects of hormones that use nucleic receptors are long-term. Signal transduction via these receptors involves little proteins, but the details of gene regulation by this method are not well understood. Nucleic receptors have DNA-binding domains containingzinc fingers and a ligand-binding domain; the zinc fingers stabilize DNA binding by holding its phosphate backbone. DNA sequences that match the receptor are usually hexameric repeats of any kind; the sequences are similar but their orientation and distance differentiate them. The ligand-binding domain is additionally responsible for dimerization of nucleic receptors prior to binding and providing structures for transactivation used for communication with the translational apparatus. Steroid receptors are a subclass of nuclear receptors located primarily within the cytosol; in the absence of steroids, they cling together in an aporeceptor complex containing chaperone orheatshock proteins (HSPs). The HSPs are necessary to activate the receptor by assisting the protein to fold in a way such that the signal sequence enabling its passage into the nucleus is accessible. Steroid receptors, on the other hand, may be repressive on gene expression when their transactivation domain is hidden; activity can be enhanced by phosphorylation of serineresidues at their N-terminal as a result of another signal transduction pathway, a process called crosstalk. Retinoic acid receptors are another subset of nuclear receptors. They can be activated by an endocrine-synthesized ligand that entered the cell by diffusion, a ligand synthesised from aprecursor like retinol brought to the cell through the bloodstream or a completely intracellularly synthesised ligand like prostaglandin. These receptors are located in the nucleus and are not accompanied by HSPs; they repress their gene by binding to their specific DNA sequence when no ligand binds to them and vice versa. Certain intracellular receptors of the immune system are cytoplasmic receptors; recently identified NOD-like receptors (NLRs) reside in the cytoplasm of some eukaryotic cells and interact with ligands using a leucine-rich repeat (LRR) motif similar to TLRs. Some of these molecules like NOD2 interact with RIP2 kinase that activates NF-B signaling, whereas others like NALP3 interact [35] [36] with inflammatory caspases and initiate processing of particular cytokines like interleukin-1. [edit]Second

messengers

First messengers are the intracellular chemical messengers (hormones, neurotransmitters, and paracrine/autocrine agents) which reach the cell from the extracellular fluid and bind to their specific receptors. Second messengers are the substances which enter the cytoplasm and act within the cell to trigger a response. Second messengers essentially serve as chemical relays from the plasma membrane to the cytoplasm, thus carrying out intracellular signal transduction. [edit]Calcium The release of calcium ions from the endoplasmic reticulum into the cytosol results in its binding to signaling proteins that are then activated; it is then sequestered in the smooth endoplasmic reticulum and the mitochondria. Two combined receptor/ion channel proteins control the transport of calcium: the InsP3-receptor that transports calcium upon interaction with inositol triphosphate on its cytosolic side and the ryanodine receptor named after the alkaloid ryanodine, similar to the InsP3 receptor but having a feedback mechanism that releases more calcium upon binding with it. The nature of calcium in the cytosol means that it is active for only a very short time, meaning its free state concentration is very low and is mostly bound to organelle molecules like calreticulin when inactive.

Calcium is used in many processes including muscle contraction, neurotransmitter release from nerve endings and cell migration. The three main pathways that lead to its activation are GPCR pathways, RTK pathways and gated ion channels; it regulates proteins either directly or by binding to an enzyme. [edit]Lipophilics Lipophilic second messenger molecules are derived from lipids residing in cellular membranes; enzymes stimulated by activated receptors activate the lipids by modifying them. Examples include diacylglycerol and ceramide, the former required for the activation of protein kinase C. [edit]Nitric

oxide

Nitric oxide (NO) acts as a second messenger because it is a free radical that can diffuse through the plasma membrane and affect nearby cells. It is synthesised from arginine and oxygen by the NO synthase and works through activation of soluble guanylyl cyclase, which when activated produces another second messenger, cGMP. NO can also act through covalent modification of proteins or their metal co-factors; some have a redox mechanism and are reversible. It is toxic in high concentrations and causes damage during stroke, but is the cause of many other functions like relaxation of blood vessels, apoptosis and erections. [edit]Redox

Signaling

In addition to nitric oxide, other electronically-activated species are also signal-transducing agents in a process called redox signaling. Examples include superoxide, hydrogen peroxide, carbon monoxide, and hydrogen sulfide. Redox signaling also includes active modulation of electronic flows [37] in semiconductive biological macromolecules

Gene-for-gene relationship
The gene-for-gene relationship was discovered by Harold Henry Flor[1][2][3][4] who was working with rust (Melampsora lini) of flax (Linum usitatissimum). Flor was the first scientist to study the genetics of both the host and parasite and to integrate them into one genetic system.[5] Gene-for-gene relationships are a widespread and very important aspect of plant disease resistance. Flor showed that the inheritance of both resistance in the host and parasite ability to cause disease is controlled by pairs of matching genes. One is a plant gene called the resistance (R) gene. The other is a parasite gene called the avirulence (Avr) gene. Plants producing a specific R gene product are resistant towards a pathogen that produces the corresponding Avr gene product. Clayton Oscar Person[6] was the first scientist to study plant pathosystem ratios rather than genetics ratios in host-parasite systems. In doing so, he discovered the differential interaction that is common to all genefor-gene relationships and that is now known as the Person differential interaction. [5]

[edit]Resistance [edit]Classes

genes

of resistance gene

There are several different classes of R Genes. The major classes are the NBS-LRR genes[7] and the cell surface pattern recognition receptors (PRR).[8] The protein products of the NBS-LRR R genes contain a nucleotide binding site (NBS) and a leucine rich repeat (LRR). The protein products of the PRRs contain extracellular, juxtamembrane, transmembrane and intracellular non-RD kinase domains.[8][9] Within the NBS-LRR class of R genes are two subclasses[7]: -

One subclass has an amino-terminal Toll/Interleukin 1 receptor homology region (TIR). This includes the N resistance gene of tobacco against tobacco mosaic virus (TMV).

The other subclass does not contain a TIR and instead has a leucine zipper region at its amino terminal.

The protein products encoded by this class of resistance gene are located within the plant cell cytoplasm. The PRR class of R genes includes the rice XA21 resistance gene that recognizes the ax21 peptide the Arabidopsis FLS2 peptide that recognizes the flg22 peptide from flagellin. There are other classes of R genes, such as the extracellular LRR class of R genes; examples include rice Xa21D [11] for resistance against Xanthomonas and the cf genes of tomato that confer resistance against Cladosporium fulvum. The Pseudomonas tomato resistance gene (Pto) belongs to a class of its own. It encodes a Ser/Thr kinase but has no LRR. It requires the presence of a linked NBS-LRR gene, prf, for activity.
[10]

and

[edit]Specificity

of resistance genes

R gene specificity (recognising certain Avr gene products) is believed to be conferred by the leucine rich repeats. LRRs are multiple, serial repeats of a motif of roughly 24 amino acids in length, with leucines or other hydrophobic residues at regular intervals. Some may also contain regularly spaced prolines and arginines.[12] LRRs are involved in protein-protein interactions, and the greatest variation amongst resistance genes occurs in the LRR domain. LRR swapping experiments between resistance genes in flax rust resulted in the specificity of the resistance gene for the avirulence gene changing.[13]

[edit]Recessive

resistance genes

Most resistance genes are autosomal dominant but there are some, most notably the mlo gene in barley, in which monogenic resistance is conferred by recessive alleles. mlo protects barley against nearly all pathovars of powdery mildew.

[edit]Avirulence

genes

The term avirulence gene remains useful as a broad term that indicates a gene that encodes any determinant of the specificity of the interaction with the host. Thus, this term can encompass some conserved microbial signatures (also called pathogen or microbe associated molecular patterns (PAMPs or MAMPs)) and pathogen effectors (e.g.,bacterial type III effectors and oomycete effectors) as well as any genes that control variation in the activity of those molecules.[10] There is no common structure between avirulence gene products. Because there would be no evolutionary advantage to a pathogen keeping a protein that only serves to have it recognised by the plant, it is believed that the products of Avr genes play an important role in virulence in genetically susceptible hosts. Unlike the MAMP or PAMP class of avr genes that are recognized by the host PRRs, the targets of bacterial effector avr proteins appear to be proteins involved in plant innate immunitysignaling, as homologues of Avr genes in animal pathogens have been shown to do this. For example, the AvrBs3 family of proteins possess DNA binding domains, nuclear localisation signalsand acidic activation domains and are believed to function by altering host cell transcription.[14