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Overview
of Cassava SSR Markers at CIAT
CIAT
has continued to lead in the development and deployment of
cassava molecular genetic tools. By developing and deploying
microsatellites or simple sequence repeat (SSR) markers, we
have built on the gains made when we published the RFLP-anchored
framework map in 1997. However, on recognizing the shortcomings
of the RFLP technology, our Unit has shifted focus toward
developing PCR-based molecular genetic tools, essentially
SSR markers. RFLP techniques are expensive, requiring the
use of hazardous radioactive probes that are not available
to many resource-poor research programs in developing countries,
and must be physically transferred from site to site under
strict safety protocols. In contrast, PCR-based markers are
robust, inexpensive to assay, easily shared among researchers,
and readily accessible in public and private domains, making
this a much more appropriate approach for developing countries.
With access to a simple text file containing the sequences
of the oligonucleotide primers for the PCR-based markers of
interest, a breeder can rapidly and efficiently evaluate the
germplasm under study.
This
technology can significantly improve the efficiency of cassava
varietal development programs and reduce time and costs by
as much as 50%, thus effectively doubling the capacity of
existing research programs. Achievements made in developing
and deploying SSR markers include:
Development
Over
500 microsatellite (SSR) markers have been developed. Many
of these have been placed on the framework map. By the end
of 2001, no less than 300 PCR-based markers were mapped.
Deployment
These
SSR markers have been made publicly available at http://www.resgen.com/products/ADDMPs.php3
and through publications.
Capacity
Building
Young
cassava scientists from Uganda, Nigeria, Ghana, Brazil, and
Ecuador have been trained in the use of cassava SSR markers.
They are now applying these markers in their work in their
respective countries.
Application
SSR
markers have been successfully applied in:
- Tagging
cassava genome loci involved in resistance to cassava mosaic
disease (CMD), a virulent disease of the crop
- QTL
mapping of the cassava genome loci controlling cassava bacterial
blight (CBB) and the cassava whitefly, an important disease
vector
- QTL
mapping of earliness and several other agronomic traits
- Mapping
of resistance to cassava green mite, a major cassava pest
- Genetic
mapping of beta carotene content
- Genety
mapping of dry metter content
Contact:
Martin Fregene
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Marker-assisted
Breeding of Resistance to Cassava Mosaic Disease
The
discovery of a qualitative and high level of resistance to
the devastating cassava mosaic disease (CMD) and molecular
markers linked to it have made marker-assisted breeding for
CMD resistance conceivable at CIAT. Progress this year includes
establishing a sexual hybridization scheme between resistant
donor lines received last year from IITA and CIAT elite parents.
Included
for genetic crosses are high carotene (precursor of vitamin
A) lines, to combine high carotene and CMD resistance, targeted
at sub-Saharan Africa. Furthermore, field experiments in Uganda
have revealed that the novel source confers resistance against
the Ugandan variant (Ug V), an aggressive recombinant strain
of the virus that caused a disease epidemic that swept through
Uganda and is now spreading into the Democratic Republic of
the Congo, Kenya, Tanzania, and Rwanda. To contain and prevent
the spread of the epidemic, a marker-assisted selection (MAS)
scheme has also been initiated to rapidly verify CMD-resistant
selections in resistance breeding. Progress is also being
made on identifying candidate genes that may mediate the molecular
basis of CMD resistance.
Contact:
Martin Fregene
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| Cassava
Transformation and Friable Embryogenic Callus Development
 At
CIAT, genetic transformation of cassava is being used as a
tool to support conventional breeding programs. The aim has
so far focused on genes (such as those for pest resistance)
not yet available in cassava germplasm, although the modification
of metabolic pathways (like starch modification and b-carotene
content) to improve cassava is also sought. Transgenic plants,
and cell lines, containing genes for pest resistance, starch
modification and herbicide tolerance, are now being produced.
Molecular tests for gene expression, and preliminary bioassays
to test efficacy of protection against cassava stemborer were
carried out this year. We observed low levels of protection.
However, more bioassays with younger larvae have to be done
to confirm observations.
Transformation
is now being achieved with friable embryogenic callus or FEC,
and using a model, and farmer preferred cultivars. We have
established FEC cell lines for four cultivars, two of which
are for the North Coast, one for the inter-Andean valleys
of Colombia, and a cultivar used as a model in transformation.
We have therefore set up a system for scaling up cassava transformation
at CIAT. Experiments are done 2-3 times a month, using Agrobacterium
or biolistics, with at least four cultivars (TMS 60444, SM
1219-9, CM 2306-4, and M Col 2215).
Contact:
Paul Chavarriaga
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Genetic
Diversity Characterized
 We
extended our study of the genetic diversity of cassava landraces,
using microsatellites, to assess the genetic diversity and
differentiation of cassava landraces from five countries in
South America, three in Central America, and four in Africa;
and also to African cassava genotypes resistant to cassava
mosaic disease (CMD). Analysis showed substantial genetic
diversity in CMD resistance, with appropriate germplasm available
for the genetic improvement of CMD resistance, as well as
other traits, particularly yield. The study of genetic diversity
in Tanzania, Nigeria, Uganda and Ghana revealed a broad and
unique diversity of local land races that can serve as the
basis for a breeding program.
The above results have been placed on the The
Cassava Molecular Diversity Network (MOLCAS) Web Site
and are available to the Cassava Community.
Contact:
Martin Fregene
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Cryopreservation
of Cassava Germplasm, Its Wild Relatives, and Cell Lines for
Tissue Culture
 More
than 10 years ago, the Biotechnology Research Unit, together
with the Genetic Resources Unit, set goals to develop methods
of cryopreservation that would lead to safer, cheaper, and
long-term conservation of genetic resources. Methods to cryopreserve
cassava germplasm were developed 4 years ago, using classic
protocols (chemical dehydration and programmed freezing).
Escobar, R.H.; Mafla, G.; Roca, W.M. 1997. A methodology for
recovering cassava plants from shoot tips maintained in liquid
nitrogen. Plant Cell Reports
16: 474 - 478. New protocols-encapsulation dehydration and
quick-freezing-have now been developed and validated with
more than 43% of the entire cassava core collection. More
than 82% of the accessions tested have recovery rates of more
than 30%, the minimum required for cryopreservation. Protocols
are now being adjusted for wild relatives of cassava, species
of which sometimes behave very poorly in vitro or even in
the field, making their conservation troublesome. Plants have
been recovered for M. esculenta ssp. flabellifolia,
M. esculenta ssp. peruviana and M. carthaginensis.
Cryopreservation
is also being used to support transformation of cassava. Developing
friable embryogenic callus cell lines is time consuming, with
the inherent risks of genetic instability and low plant recovery
over time. Cryopreserving FEC cell lines is therefore a viable
alternative. FEC
cell lines of two cassava cultivars (TMS 60444 and M Col 2215)
have been frozen and recovered. Because transformation of
cassava requires the development of FEC for each specific
cultivar, we expect to build up a cryopreservation bank of
FEC cell lines of various cassava cultivars.
Contact:
Roosevelt Escobar
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