Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
1999-08-31
2003-04-08
Yucel, Remy (Department: 1636)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C435S320100, C536S024100
Reexamination Certificate
active
06544733
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to a new method to rapidly identify genes that function in non-host resistance. It also relates to genes identified by this method that enhance levels of disease resistance if expressed in susceptible plants.
BACKGROUND OF THE INVENTION
Genetic diversity is an important factor in the balanced evolution between plants and their pathogens. In natural systems, outbreeding plant populations interact with mixed pathogen populations. This interaction is often dependent on the presence of resistance (R-) genes in the plant and avirulence (avr) genes in the pathogen. The outbreeding plants share pools of R-genes, and the plant pathogens produce a variety of elicitors, directly or indirectly produced by the avr genes. Individual plants that contain R-genes that somehow recognize one of the elicitors produced by an infecting pathogen are resistant against this pathogen.
R-gene mediated resistance usually results in a hypersensitive response (HR), observed as rapid necrosis at the infection site. Apparently, the activated R-gene triggers a signal transduction event leading to apoptotic cell death, which may prevent the invading pathogen from spreading beyond the infection site and trigger resistance in non-infected adjacent cells.
Over the last five years, a number of R-genes have been cloned. The most ubiquitous class of R-genes encode proteins with a C-terminal leucine rich repeat, an N-terminal nucleotide binding site, and a conserved stretch of amino acids with the consensus sequence GLPLAL. Examples of this class of R-genes are Rps2 (Bent et al., 1994), N (Whitham et al., 1994), L6 (Lawrence et al., 1995), M (Anderson et al., 1997), and Rpm
1
(Grant et al., 1995). Progress has also been made in the identification of proteins involved in R-gene mediated signal transduction. Recent papers report the involvement of protein kinases, putative transcription factors, and lipase-like proteins in R-gene signalling (reviewed by Innes, 1998). Recently, it has been shown that the engineering of these signaling components may also lead to enhanced levels of disease control in plants (Cao et al., 1998).
It is believed that R-genes do not provide protection against all genotypes of a pathogen, i.e., pathogens within a species do not all produce the same elicitor. It is therefore likely that infections of outbreeding populations will result in the survival of part of the population only. Modem agriculture may likely disturb the balance between plants and pathogens. Outbreaks of a disease that several decades ago would impact a relatively limited number of plants can now cause devastating epidemics.
To prevent major losses to diseases, plant breeders attempt to introgress resistance against the most important pathogen races into elite cultivars. In most cases, this is a never-ending battle because resistance against one or several genotypes of a pathogen will select for occurrence of other genotypes. For example, the subsequent introgression of eleven R-genes from the resistant wild potato species
Solanum demissum
into cultivated susceptible potato cultivars resulted in all cases in the emergence of virulent genotypes of the pathogen
Phytophthora infestans
. Classical breeding is by definition based on crossing programs and, therefore, can only transfer resistance traits between different accessions or cultivars of the same plant species or between plant species that are sexually compatible. This resistance is often referred to as “host” resistance. Temporal control of many pathogens including the following have been obtained by introgression of R-genes:
Phytophthora infestans, Phytophthora megasperna, Puccinia graminis, Puccinia recondita, Puccinia sorghi, Puccinia coronata, Puccinia helianthi, Puccinia striiformis, Erysiphe graminis, Ustilago hordei, Ustilago avenae, Uromyces phaseoli, Peronospora farinosa, Pseudomonas syringae, Xanthomonas oryzae, Cladosporium fulvum
, brown plant hopper, aphids, hessian fly, and tobacco mosaic virus.
A few R-genes have been identified that provide resistance against most races of a particular pathogen. Of particular interest are the rice Xa21 gene that controls most races of
Xanthomoas oryzae
(Mazzola et al., 1994; Song et al., 1995), the wheat Lr34 gene involved in resistance to most leaf rusts, and the barley Rpg1 gene that protects plants against almost all stem rusts. However, these R-genes are rare and may be broken by new aggressive races.
A superior source of resistance that provides broad-spectrum and durable disease control but is unaccessible to classical breeding is the so-called “non-host” resistance. A plant species displays non-host resistance if all sexually compatible accessions and cultivars of that particular species or very related species are resistant to all genotypes of a particular pathogen. Due to the lack of susceptible material within those plant species, it is impossible to determine the genetic basis of non-host resistance.
To date, no genes have been cloned that are known to be involved in active non-host resistance. However, it is possible that such genes resemble the R-genes isolated from sources displaying host resistance. Support for this hypothesis comes from studies on the interaction between
P. infestans
and the non-host plant species
Nicotiana tabacum
(tobacco). The resistance of tobacco correlates with its ability to respond with an HR to infection, suggesting that the resistance of tobacco against
P. infestans
is based on an active defense mechanism controlled by R-genes (Kamoun et al., 1997). Thus, the non-host resistance of tobacco appears to be “active”, and is different from “passive” resistance that is based on factors such as the presence of preformed pathogen inhibitors or the absence of factors that are essential for pathogen growth (Ride, 1985).
It can be envisioned that expression of certain cloned non-host resistance genes in susceptible crops would provide the broad-spectrum and durable disease resistance levels that are needed in modern agriculture. However, it is impossible to isolate non-host resistance genes through genetics-based methods. Here, the inventors have developed a new technique based on the isolation and screening of large numbers of genes that are associated with active non-host resistance. The screening is performed in plants that are both susceptible to certain target pathogens and highly accessible to transformation. By implementing this technique, a number of genes have been identified that enhance, or are expected to enhance, levels of disease resistance if expressed in susceptible plants.
SUMMARY OF THE INVENTION
The present invention relates to a method to screen genes associated with non-host resistance for those genes that enhance levels of resistance if expressed in susceptible plants, by transforming tissue of a pathogen-susceptible plant with these genes, challenging the transformed tissue with a pathogen or its elicitor, and observing enhanced defense and/or HR responses. In a particular embodiment of the invention, homologs of R-genes from tobacco are identified by gene amplification, cotransformed with the INF1 elicitor of
Phytophthora infestans
into leaves of
Nicotiana benthamiana
, and screened for the presence of a hypersensitive response, which indicates functionality. In another embodiment, genes associated with non-host resistance are identified by first selecting genes that are induced by target pathogens in the non-host but not (or not as much) in susceptible hosts, and second screening them for their ability to enhance resistance against a model pathogen such as the bacterial pathogen
Pseudomonas tabaci
if transiently overexpressed in leaves of
N. benthamiana
plants.
In one aspect, the present invention provides novel nucleic acid sequences (SEQ ID NO:57 and SEQ ID NO: 1-10 and SEQ ID NO:58, 60, 62) that can confer disease resistance to
Phytophthora infestans
to plants. A further embodiment of the invention provides novel protein sequences (SEQ ID NO: 11-20 and SEQ ID NO:59) involved in disease re
Rommens Caius M. T.
Swords Kathleen M M
Yan Hua
Zhang Bei
Ball Timothy K.
Hoerner, Jr. Dennis R.
Katcheves Konstantina
Monsanto Technology LLC
Yucel Remy
LandOfFree
Method of identifying non-host plant disease resistance genes does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Method of identifying non-host plant disease resistance genes, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Method of identifying non-host plant disease resistance genes will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3110264