Biotechnology, Breeding and Seed Systems for African Crops
Biotechnology
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Molecular
characterization of a novel stress-inducible antioxidant gene, XvPer1,
from the resurrection plant Xerophyta
viscosa Baker. S.B. Mowla,
J.A. Thomson, J.M. Farrant, S. G. Mundree Environmental stresses occur in various parts of the
world, especially in response to global climate change. As a result,
crop plant productivity is negatively affected globally, but more so in
sub-Saharan Africa. The presence of high light intensities often occurs
in addition to water limitation and can be extremely damaging to
photosynthetically active tissues leading to the formation of oxygen
free radicals. If unquenched, these molecules cause considerable damage
to the subcellular milieu. Reactive oxygen species (ROS) play a
significant role in causing damage to living cells under severe stress
conditions. To combat oxidative stress, complex protective mechanisms
have been evolved by plants to mitigate and repair the damage initiated
by free-radicals. By differential screening of a X. viscosa cDNA library, genes were identified that were upregulated
when the plant was in a dehydrated state. XvPer1 was one of them, coding
for a novel antioxidant enzyme. A
cDNA named XvPer1
was isolated from X.
viscosa by differential screening of a cDNA library.
Characterization of the cDNA showed that XvPer1 has an ORF of 849 bp encoding a polypeptide of 210 residues.
The XvPer1 cDNA contains a
putative polyadenylation site and codes for a putative bipartite nuclear
localization signal (NLS) near the 3’-end. The cDNA corresponds to
1-Cys peroxiredoxin, an evolutionary conserved thiol-specific
antioxidant enzyme. XvPer1 polypeptide shows significant sequence
identity (~70%) to other recently identified plant 1-Cys peroxiredoxins.
The amino acid sequence proposed to constitute the active site of the
enzyme (PVCTTE) is highly conserved in XvPer1. Southern blot analysis
revealed a single copy of XvPer1
in the genome. Analysis of the gene expression using northern blot
indicated that the transcript is absent in fully hydrated X. viscosa tissues but is induced under abiotic stresses such as
dehydration, heat (42oC), high light (1500 mmol
m-2 s-1) and when treated with abscisic acid (100 mM)
and sodium chloride (150 mM). Western blot data correlated with the
patterns of expression of XvPer1.
Preliminary localization studies using immunofluorescence revealed that
XvPer1 is localized in the nucleus of dehydrated X.
viscosa leaf cells. These results suggest that the stress-inducible
gene Xvper1 may function to
protect nucleic acids within the nucleus against oxidative injury. Novel
determinants of osmotic stress tolerance using the monocotyledonous
resurrection plant Xerophyta
viscosa S.G. Mundree
& J.A.Thomson Water
has become a major limiting factor in world agriculture.
In general, most crop plants are highly sensitive to even mild
dehydration stress. There are, however, a few genera of plants unique to
sourthern Africa called “resurrection plants” which can tolerate
extreme water loss or desiccation. The unique ability of these plants to
withstand severe water loss (greater than 90%) makes them an ideal
system to study osmotic stress tolerance. Xerophyta
viscosa, a representative of the monocotyledonous resurrection
plants, was used to isolate genes that are functionally important in
osmotic stress tolerance, and those that are differentially expressed
during dehydration stress, respectively.
A cDNA library, based on mRNA isolated during various stages of
dehydration was inserted in Lambda Phage Zap II vector (Stratagene).
The advantage of using the above vector is that it allowed for
the rescue of phagemids containing the cDNA inserts by helper
phage-mediated in vivo
excission. The rescued
phagemids were used to infect a specific strain of Escherichia
coli (srl: Tn 10 mutant) which was grown under severe osmotic
stress. With IPTG present
in the medium, bacterial cells containing cDNA that conferred osmo-tolerance,
emerged as colonies on minimal medium plates supplemented with 1.2 M
sorbitol. Nine colonies containing cDNA which conferred functional
sufficiency to the osmotically-stressed bacterial cells were identified.
Two of the cDNA have been characterised extensively viz. ALDRXV4,
an aldose reductase homologue, and XVSAP1, a stress-associated protein.
The second strategy involved the differential screening of 192 randomly
selected cDNA clones from above cDNA library. Of the 192 cDNA screened,
30 were found to be up-regulated. cDNA
have been sequenced and characterised.
The identities of these proteins included: XCDH, a dehydrin;
XVLEA, a lea-like homogue; XVGS, galactinol synthase, XVCBP, EF-hand
calcium-binding protein; XVHSP90, a heat shock 90 protein; and XVPER, a
peroxiredoxin. All six of
the cDNA have been found to be expressed during dehydration streess in
the resurrection plant and they have also been implicated in other
stresses, particularly those that have an osmotic stress component.
Evaluation of cassava clones for yield, farmer acceptability and disease
and pest resistance Evaluation
of cowpea aphid-borne mosaic virus (CABMV) coat protein and RNA mediated
resistance strategies in Nicotiana
benthamiana R. Mundembe
& I. Sithole-Niang Pathogen-derived resistance to viruses has been very
successful for many virus groups, but has not been widely used to
control viral diseases of importance to farmers in developing countries.
Cowpea aphid-borne mosaic potyvirus (CABMV) is one of the
pathogens that significantly reduces cowpea yields of resource poor
farmers. The goal of this
study was to determine the most effective virus resistance mechanism
using the CABMV coat protein (CP) gene.
The CP gene was amplified using the reverse transcription-polymerase
chain reaction (RT-PCR) technique, incorporating an ATG translation
start codon and the Kozak consensus sequence for optimum expression in
plants, and cloned into a PCR cloning vector pCRII, to result in a
plasmid that was named pCRII-CPk. The CP gene fragment was excised from pCRII-CPk using Bam
HI and Sal I and cloned into a similarly digested expression cassette
vector pCa2Nos, between the promoter and terminator. The plasmid pCRII-CPk was also used as a template to amplify
an untranslatable CP gene with stop codons in all three reading frames (CPstop),
the CP gene in an anti-sense orientation (PC), and the central region of
the CP gene (CPcore). All
three amplification products were cloned into pCa2Nos, between the
promoter and terminator, and the entire insert was ligated into the
unique Hind III site of the
binary plasmid pBI 121, to result in plasmids pBI121-CPk, pBI121-CPstop,
pBI121-PC and pBI121-CPcore. The
constructs were used to transform Agrobacterium tumefaciens which was then co-cultivated with Nicotiana
benthamiana leaf sections. The
regenerated plants were analyzed by PCR and Southern blotting, and the
seeds were germinated on kanamycin and challenged with CAMBV-infected
sap, viral RNA and whole virions. A
search for random amplified polymorphic DNA (RAPD) and simple sequence
repeat (SSR) markers that are linked to Striga
asiatica resistance in sorghum
C.S.
Mutengwa,
P. Tongoona, Z. Dhlamini & I. Sithole-Niang
Striga asiatica (L.) Kuntze, or witchweed, causes major yield
reductions in principal cereal grain crops of Zimbabwe, including
sorghum [Sorghum bicolor (L.)
Moench]. Yield losses can be up to 100% in heavily infested fields. The
development of high yielding cultivars with durable resistance to Striga (Striga spp) is the
most feasible method for achieving sustainable control of this parasite
in smallholder agriculture. Development of Striga
resistant cultivars by conventional breeding is however slow and has
been hampered by the lack of efficient and reliable screening techniques
in breeding programs. Identification of molecular markers that are
linked to witchweed resistance can expedite the development of resistant
cultivars through adoption of appropriate marker assisted selection
(MAS) strategies. We therefore sought to identify molecular markers that
are linked to S. asiatica seed germination stimulant production, which is one the
recognised mechanisms of resistance to witchweeds. A segregating F2
population derived from crosses between cultivars SV-1 (high germination
stimulant producer, Striga-susceptible)
and SAR 29 (low germination stimulant producer, Striga-resistant)
was used for this purpose. Parental and F2 genotypes were screened for S.
asistica seed germination stimulant production using the agar gel
technique (AGT). The index of resistance was the maximum distance from
the sorghum root at which S. asiatica seeds were found germinating. Genotypes with a
germination distance of less than 1 cm were classified as resistant and
those with a distance greater than or equal to 1 cm were classified as
susceptible. DNA of AGT-classified genotypes were then assayed using
random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR)
markers. Linkage analysis was performed using the computer software
MAPMAKER 3.0b, at a threshold LOD score of 3.0 and a 0.50 maximum
recombination frequency. Segregation
ratios of high to low F2 stimulant producers did not differ
significantly (P £
0.05) from the expected ratio of 3:1. It was therefore concluded that a
single recessive gene controls low witchweed seed germination stimulant
production in cultivar SAR 29. A total of 68 marker loci (10 SSR and 58 RAPD) were scored in the
segregating F2 population. Segregation analysis of individual marker
loci using the chi-square goodness-of-fit test revealed that 43 (62.32%)
of the loci segregated normally while 26 (37.68%; 2 SSR and 24 RAPD)
exhibited distorted segregation (P £
0.05). Cosegregation analysis resulted in 39 markers (3 SSR and 36 RAPD)
being assigned to nine linkage groups spanning a total distance of 817.1
cM. Thirty loci, including the locus for low stimulant production, were
not assigned to any linkage group. The map was constructed from an
average of 60 individuals per primer. The average, minimal and maximal
distances between markers were 11.8 cM, 3.2 cM and 176.1 cM,
respectively. More RAPD and SSR markers are currently being added to the
molecular linkage map. This will improve coverage of the genome and
facilitate identification of molecular markers for S.
asiatica resistance in sorghum.
Development
of highland banana cell suspension system: A critical stage in genetic
improvement of the banana
P. Namanya,
G. Mutumba, S.M. Magambo, W. Tushemereirwe The East African highland banana, the most important
staple food in Uganda, is susceptible to a range of pests and diseases.
This has been attributed to a limited genetic variation of the banana
group. Development of resistant genotypes using a genetic transformation
approach has been identified as a potential option to explore. The
initial objective was to establish a system to generate embryogenic cell
suspensions for East African highland bananas. Cell suspensions are the
material of choice for genetic transformation of banana because of their
regeneration capacity through somatic embryogenesis. Immature male
flowers were used as explant source.
Embryogenic callus with somatic embryos and friable tissue was
obtained after a culture period of 4 months on callus induction medium
designated M1. Subsequent
cell suspensions were initiated and maintained in liquid medium MA2
(Cote et al., 1996). The cells
in suspension were embryogenic and a cell growth rate of 1.5-2.0 scv per
month was observed. The embryogenic cell suspension stage is critical
for introduction of genes of resistance into the bananas. Regeneration produced up to 2.18x102 embryos per
0.05-0.1ml scv. Germination of these embryos was observed at 2.78% and
3.27% for two cell suspension lines. Plantlet recovery was 60-100%, all
being normal, plants with root and shoot at weaning. This study clears
the key hurdle to application of genetic improvement of East African
highland bananas using genetic transformation approaches. Utilization of molecular markers to characterize and map useful genes of the cultivated African rice Oryza glaberrima M.N.
Ndjiondjop,
A. Alonge, S. Mc Couch
& M. P. Jones This
paper summarizes the progress made by the West African Rice Development
Association (WARDA) in developing the genetic tools and plant material
required for the identification and characterization of valuable genes
from Oryza glaberrima, and to assist the breeders by using
molecular-markers in the selection of genotypes that combine the high yield potential of O.
sativa with the stress tolerance of
O. glaberrima.
WARDA used anther culture and backcrossing to produce
several hundred fixed lines from interspecific crosses between varieties
CG14 (O. glaberrima) and
WAB56-104 (O. sativa, subsp. japonica).
We selected 50 doubled haploid and 50 backcross lines with superior
adaptation in farmer field. A set of 131 polymorphic microsatellites
dispersed over the 12 chromosomes of rice was used to assess the proportion of the genome from each parent in the interspecific lines. From
the graphical genotyping of all lines, we observed the
introgression of O. glaberrima alleles at the end of the long
arm of chromosome 2, the short arm of chromosome 6 and in the middle of
chromosomes 8, 9 and 10. The
introgressed fragment on chromosome 6 is probably linked to the
gamete-killer gene reported on this chromosome; however, on chromosomes
2, 8, 9 and 10 mechanisms other than the sterility gene may be involved.
On average, the introgressed segments from O. glaberrima
were small (average size 5 cM) and distributed on almost all
chromosomes, indicating that recombination occurs frequently in this
interspecific combination and that there is no genome-wide barrier to
recombination during meiosis. The mean of O. glaberrima alleles
across lines was 12% and the
mean of O. sativa alleles 86%. Some non-parental alleles were
identified. We believe that these alleles, which are not linked to the
high mutation rate due to microsatellites, are due to out-crossing and
have played an important role in increasing the genetic diversity of the
lines. In addition, the lines
were fixed in all parts of the genome. The phenotyping and
graphical genotyping of these lines has enabled the selection of three
interspecific lines. They are used to develop the near isogenic material
necessary to identify genes/QTLs controlling characters, such as wide
droopy leaves, number of grains per panicle, panicle size, tolerance to
blast and drought. These lines have also provided the opportunity to
characterize and to assess the genetic basis of male sterility.
To establish the marker-assisted
selection system at WARDA, we developed a genetic mapping program. Advanced
backcross and dihaploidization strategies from BC2F1
lines were used to produce segregating materials.
A good source of single gene resistance
to RYMV, a major constraint to rice production in several countries in
West Africa, has been identified in the variety Gigante (O.
sativa, subsp
indica) and mapped on chromosome 4. Fine
mapping allowed the determination of three microsatellite markers close
to the RYMV-resistance gene (2 cM), providing the opportunity to easily
transfer the resistance gene from Gigante into other genetic backgrounds
using marker-assisted selection. Three elite rice varieties, namely
BG90-2, Bouaké189 and Jaya have been selected by WARDA, since
they are well adapted to African lowland conditions, but these are
highly susceptible to RYMV. Two backcross progenies have been produced
and evaluated for resistance to RYMV. The BC2F1
resistant materials are now available and are being used to continue the
backcrossing process. Gene flow
between cultivated and wild cowpea in Kenya R.S. Pasquet,
A. Ochieng & K. Thoen The wild progenitor of cultivated cowpea (Vigna
unguiculata ssp. unguiculata var. spontanea) is encountered in all
humid lowland ecologies of Kenya, including the areas surrounding Lake
Victoria, between 1000 and 1500 m in Central Kenya, and in the coastal
plains. In the coastal plains, a second wild cowpea subspecies (V.
unguiculata ssp. pubescens) is also encountered but hybridization
between this subspecies and cultivated cowpea is not common. In all
these areas, wild cowpea shows a weedy behavior and is much more present
in disturbed areas, often in the vicinity of cultivated cowpea.
Genetic structure of representative populations as well as
outcrossing rates (in natural conditions) are assessed using isozyme
markers. Results suggest that gene flow between populations is low.
Within populations, isozyme polymorphism is lower than expected (since
some populations have an outcrossing breeding system) and may be too low
to accurately really assess gene flow events.
Floral biology data, i.e. floral scent, nectar secretion, number
of pollen grains and ovules, anthesis time, stigma receptivity, the
anthers and stigma relative position of, and seed set are presented.
Breeding systems range from total outcross (few wild populations) to
inbred (in cultivated plants). Pollinators
are identified as carpenter bees and several Megachiles. They collect
nectar and are normally visit each flower twice (at sunrise and once
after the nectar is refilled). They do not exhibit flower color
preferences or species prefences (when V.
reticulata flowers are close to V.
unguiculata flowers). They fly short distances while foraging and up
to now, attempts to prove gene flow between plants beyond 50 m have
failed. Fitness of hybrids
is assessed. F1 hybrids are obiously more fit than their parents,
especially when one mother plant is a cultivated plants. They produce
many seeds. However, these seeds are not dormant which may explain the
low amount of hybrid plants found in natural populations. Allelochemical
mechanisms of Striga hermonthica
suppression by Desmodium uncinatum J.A. Pickett,
Z.R. Khan, A. Hassanali, W. Overholt, T.M. Khamis, A.M. Hooper, L.J. Wadhams
& C.M. Woodcock During investigations into the control of insect
damage to maize crops in subsistence farming in Kenya, which involved
intercropping with repellent plants, the fodder legumes silverleaf (Desmodium
uncinatum) and greenleaf (D.
intortum) were also found to reduce dramatically the infestation of
maize by parasitic witchweeds such as Striga
hermonthica. This
effect was confirmed by further field testing and shown to be
significantly greater than that observed with other legumes, e.g.
cowpea, as were the concomitant yield increases. The mechanism was
investigated, and although soil shading and addition of nitrogen
fertiliser showed some benefits against S.
hermonthica infestation, a putative allelopathic mechanism for D.
uncinatum was also observed. In screenhouse studies, a highly
significant reduction in S. hermonthica infestation was obtained when an aqueous solution,
eluting from pots in which D.
uncinatum plants were growing, was used to irrigate pots of maize
planted in soil seeded with high levels of S.
hermonthica. Growth of
the parasitic weed was almost completely suppressed, whereas extensive
infestation occurred with the control eluate. Laboratory investigations
into the allelopathic effect of
D. uncinatum, using samples of
water-soluble chemical components exuded from cleaned roots,
demonstrated that this involved a germination stimulant for S.
hermonthica and also an inhibitor for haustorial development. Marker-assisted
selection for maize improvement: practical examples and strategies J.M. Ribaut,
M. Banziger, K. Pixley & D. Hoisington The application of molecular markers to plant
breeding can be divided into three main categories: (1) the
characterization of germplasm, known as fingerprinting; (2) the genetic
dissection of the target trait, (actually the identification and
characterization of genomic regions involved in the expression of the
target trait); and, (3) following the identification of the genomic
regions of interest, crop improvement through marker-assisted selection
(MAS). The first two applications have proven their value by generating
knowledge about the genetic diversity of germplasm, thereby allowing
placement into heterotic groups and a better understanding of the
genetic basis of agronomic traits of interest. Given the results already produced in maize at the molecular level,
including QTL analysis, gene cloning, expressed sequence tag (EST)
identification, and the development of new molecular applications, MAS
for maize improvement is becoming more and more efficient. For simply
inherited traits – (those that have high heritability and are
regulated by only a few genes-) the use of molecular markers to
accelerate germplasm improvement has been well documented. Such work has
proven successful by (1) tracing favorable alleles in the genomic
background of genotypes of interest; and, (2) identifying individual
plants in large segregating populations that carry
the favorable alleles. In the case of line conversion – (the
transfer of elite alleles at one or several loci from a donor to a
recipient line through backcrosses)- results of simulations conducted at
CIMMYT demonstrate the importance of the selectable population size.
Population size is essentially dependent on the number of genotypes
heterozygous at the target loci involved into the selection and this
parameter is the first one to be considered when designing a MAS
experiment. For concomitant allelic introgression from a donor line into
a large number of recipient lines, MAS at unselected loci is conducted
only once at an advanced BC cycle, while MAS at selected loci is
conducted at each cycle. MAS
for polygenic trait improvement is still in an exploratory phase, with
only a few successful experiments reported. After crossing elite
material for a target environment (good by good), innovative experiments
considering MAS at the early stage of recombination and for one or a few
cycles are ongoing at CIMMYT. Strategy development for polygenic trait
improvement through molecular markers is a very dynamic area of
investigation, because optimal strategies evolve together with the
genetic information provided by ongoing and emerging technology.
Considering the new type of information provided at the gene expression
level, it is also time to think about new conventional breeding
strategies to better complement molecular and conventional approaches.
To illustrate our ideas on MAS strategies, results for QPM (cloned
gene), MSV resistance (major QTL) and drought tolerance (polygenic
trait) will be presented in details as case studies of MAS experiments
conducted at CIMMYT. Review of
tissue culture and genetic studies of East African highland bananas in
Uganda P.R. Rubaihayo,
S. Kassim, S. Nanteza & A.K. Tugume Banana is grown in Uganda by over 1.8 million farmers
on 1.5 million hectares of land, typically on small (0.5 ha) stands.
A decline and sometimes disappearance of banana cultivation,
attributed largely due to increasing pests and diseases of highland
bananas, has been witnessed
in the last three decades. In
order to address this situation it was necessary in the first instance
to provide farmers with suckers free from non-obscure pests and
pathogens that are transmittable in contaminated planting materials
through conventional methods of propagation. Available control measures
of these pests and diseases are often beyond the means of most farmers.
However, host resistance would check further losses due to these
constraints. Due to
difficulties associated with the conventional breeding of Musa, which includes lack of useful genetic variability and low
levels of female fertility, efforts have been made to develop tissue
and/or cell culture protocols to help expose the tissue and/or cells to
mutagenic agents and/or transformation systems so as to create
variability or transfer resistance genes to the East African Highland
bananas [EAHB]. PCR-based
fingerprinting technique (AFLP) has been used to establish the genetic
relationships among EAHB clones. In
vitro propagation protocol using shoot-tips has been established for
EAHB cultivars and studies of production and regeneration of somatic
embryos in EAHB initiated. The genetic relatedness of the EAHB has been
determined and found to be very close, indicating that these bananas
probably originated from a single clone or very closely related clones.
The work reported complements the classical breeding methods of
improvement going on in the country aimed at producing banana cultivar
resistant to the major pests and diseases. Resistance to African cassava mosaic virus conferred by a mutant of the
putative NTP-binding domain of the rep gene (AC1) in Nicotiana benthamiana A. Sangaré We
constructed a mutation in DNA-A of African Cassava Mosaic Virus (ACMV)
to alter the putative NTP-binding domain site in the
replication-associated protein gene (AC1). When transgenic Nicotiana
benthamiana plants expressing the mutated gene were infected with
ACMV, the plants exhibited tolerance to infection, consisting in a delay
in symptom appearance and/or the presence of mild symptoms. In addition,
the resistant plants accumulated less viral DNA than non-transgenic
palnts. As judged by northern blot analysis and symptom development of
segregating progeny from different lines, a high level of expression of
the mutant is essential for the development of resistance. Issues
related to the use of different versions of AC1 for the control of ACMV
are discussed. Can wild relatives of maize provide new sources of
resistance to the root hemi-parasite Striga
hermonthica? J.D. Scholes, A.L. Gurney, D.
Grimanelli, D. Hoisington & M.C. Press The
haustorium is an organ that forms a physiological bridge between host
and parasite acting as a conduit for host derived solutes. The root
hemiparasite Striga is
dependent on its host for water, nitrogen and some carbon, however, the
role of the haustorium in solute acquisition and metabolism is
relatively unknown. Studies of other parasitic plants such as mistletoes
suggest that the haustorium may play an important role in the metabolism
of host-derived nutrients. To date, complete resistance to Striga infection has not been identified in maize, however, a near
relative of maize, Tripsacum
dactyloides, did not support Striga
growth. Histological analysis demonstrated inferior haustorial
development on Tripsacum
compared with haustorial development on maize, in particular poor tissue
differentiation. The hyaline body, a
metabolically active region of cells within the haustorium, was either
absent or poorly developed. A lack of internal strucure is likely
to affect haustorial function which may have serious consequences for
parasite nutrition and subsequent development.
Non-functional haustoria on Tripsacum
may be a result of the absence of a positive signal from the host or the
presence of a negative signal. Secondary haustoria produced by Striga
were manipulated so that one individual Striga
plant could be attached to two hosts; a susceptible maize host and a
resistant Tripsacum host. Our
aim was to determine whether i) a positive signal(s) produced by maize
could influence haustorial development on Tripsacum
or ii) whether a negative signal produced by Tripsacum
could influence haustorial development on maize. Early results strongly
suggest that Tripsacum may
produce a negative signal influencing haustorial development and
preventing successful parasite infestation. Development
of an electrotransformation protocol for cowpea (Vigna
unguiculata) I. Sithole-Niang,
R. Mundembe & R. Allison Cowpea is a major source of dietary protein in most
African diets. It suffers
from a wide range of production constraints including viral diseases.
Cowpea aphid-borne mosaic potyvirus (CABMV) is the most important
viral pathogen of cowpea in major cowpea growing regions of the world.
While breeding for resistance is an ongoing activity is various
laboratories, the availability of resistant germplasm is limited.
Our strategy has been to use pathogen derived resistance to
introduce viral genes into cowpea.
However, this effort has met with limited success due to the lack
of an efficient, reliable and reproducible transformation and
regeneration system. Our present goal is to attempt the transformation
of cowpea seedlings using electrophoresis.
The coat protein gene of CABMV was cloned into the binary vector
pBI 121 in various forms designed to confer coat protein-mediated
resistance and sense- and antisense- RNA mediated resistances.
The DNA constructs were electrophoresed into the apical meristem
of developing cowpea seedlings under various conditions including
different voltage and current settings as well as pretreatment of
seedlings with acid or plant growth regulators.
Preliminary screening was done using GUS assays, and the plants
allowed to set seed. The
seed was harvested and will be replanted in-vitro on kanamycin. Further
analysis will be done using the polymerase chain reaction (PCR) and
Southern blot hybridization to determine the most effective regimen. Biotechnology
applications in breeding for Striga
asiatica resistance at the University of Zimbabwe P.B. Tongoona,
C.S. Mutengwa, C. Musimwa, I. Sithole-Niang & A. Matibiri Biotechnology has the potential to revolutionise the
breeding of crop cultivars because of the potential speed and precision
of selection for desirable genotypes. One aspect of biotechnology that
can be used in crop breeding is marker-assisted selection (MAS), which
involves indirect selection for a trait of interest through molecular
markers that ‘tag’ single genes or quantitative trait loci (QTL).
At the University of Zimbabwe, we have chosen Striga asiatica resistance in Sorghum
bicolor as an ideal character for application of MAS because field
screening is difficult and imprecise, S.
asiatica resistance is simply inherited and controlled by a single
recessive gene, and no resistant, high yielding sorghum cultivars have
been developed to date. Random
amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR)
markers were used to identify markers that are linked to S.
asiatica resistance in an F2 sorghum population derived from a cross
between a susceptible and a resistant parent. Details of this work are
to be presented at this meeting. We also sought to study molecular
diversity of S. asiatica
populations collected from different regions of the country and from
different host plants. This was done because differential responses of
different cereal crops have been noted when they are grown in different
geographical regions. RAPD analysis of S.
asiatica populations collected from different geographical regions
of Zimbabwe and different host plants revealed significant diversity.
The 24 populations that were used in this study clustered in five
groups. One of the groups contained accessions from the same
geographical region while the other groups contained accessions from
different regions. This made it difficult to make generalizations on the
regional basis of the relationships. It is noteworthy that different
biotypes were identified in this study. However, it is not known if
these biotypes differ in their virulence to different cultivars and host
cereals. Further investigations are required to elucidate this issue.
The presence of these biotypes apparently indicated the need for multi-locational
testing of any cultivars developed for resistance against S.
asiatica. Distribution of
angular leaf spot of common bean in Kenya and virulence diversity of the
causal agent, Phaeoisariopsis
griseola (Sacc.)
Ferr. I.N. Wagara,
A.W. Mwangómbe, J.W. Kimenju & R.A. Buruchara Angular
leaf spot, caused by Phaeoisariopsis
griseola (Sacc.) Ferr., is among the most important diseases of
common bean (Phaseolus vulgaris)
in Kenya, causing yield losses of up to 80%. When weather conditions are
favourable, the disease can be very destructive with most crop losses
resulting from premature defoliation. Breeding for resistance to this
disease is complicated by wide pathogenic variation within the causal
agent. A survey
conducted to determine prevalence, incidence and severity of angular
leaf spot revealed that the disease is widely distributed in Kenya. The
disease was prevalent in all the five districts surveyed and disease
incidence and severity were generally high. Fifty-three percent of the
farms visited had mean disease incidence of more than 50%, and 29% of
the farms had average severity scores of over 25%. P.
griseola was isolated from infected bean leaves collected from
randomly selected fields and the extent of virulence diversity in the
pathogen population determined by inoculating 12 bean varieties. Based
on their virulence reactions on the differential varieties, 100 isolates
of the pathogen were grouped into 44 races. Eight of the races
exclusively infected large-seeded bean varieties and were therefore
categorised as Andean. Twenty-nine races were of the Mesoamerican group
that infects both the large- and small-seeded varieties, and 7 races
belonged to the Afro-Andean group. These results indicate a wide
pathogenic variability in the angular leaf spot pathogen that must be
taken into consideration when developing and selecting bean varieties
with resistance to P. griseola.
Preliminary
results on surveys of banana streak badnavirus (BSV) in Kenya A.
Wangai & L. Karanja Banana streak virus (BSV) is one of the most widely distributed viruses of plantains and bananas. It has been reported in more than 40 countries where bananas are grown. Up to until 1997, the disease was not reported in Kenya, although it has been known to occur within the East African region (Uganda, Tanzania, Rwanda and Zanzibar) since 1994. The aim of this study was to re-evaluate the current status of the disease in the country The disease evaluation was done by carrying out a disease diagnostic survey covering the major banana growing regions in Kenya in Western, Central, Eastern and Coastal regions. Field BSV symptoms were observed in the crop, including chlorotic streaks, leaf deformation, stunting, cigar leaf death, distortion of peducle, bunch or fruits, and internal pseudostem necrosis. The virus characteristics, occurrence and symptom indexing were determined by serological tests ( ELISA.-(DAS) and PCR techniques. |
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