African Crop Science Conference Proceedings, Vol. 9. pp.

289 - 291
Printed in Uganda. All rights reserved
ISSN 1023-070X/2009 $ 4.00
© 2009, African Crop Science Society

Seed regeneration practices at the Southern African Development

Cooperation (SADC) Plant Genetic Resources Centre
L. LEROTHOLI QHOBELA & W.Y.F. MARANDU
SADC-Plant Genetic Resources Centre, Private/Bag CH6, ZA15302, Great East Road, Chalimbaba, Lusaka, Zambia
Bioversity International, Programme on Understanding and Managing Biodiversity. c/o AVRDC-RCA,
P.O. Box 10 Duluti, Arusha, Tanzania

Abstract Seed genebanks provide the most practical method for preserving large amounts of genetic material within a small space.
Seeds in genebanks, eventually, deteriorate whatever the conservation conditions. Genebanks managers are, therefore, faced with the
challenge of ensuring that samples stored are genetically very similar to the original sample collected in the field. Low temperature
and optimizing seed moisture content delays seed deterioration, but, when seed viability decreases to below 85%, regeneration is
recommended. Regeneration process appears routine and necessary but it may lead to losses of valuable traits if sufficient attention
is not given to factors affecting genetic integrity of the accession. Genebank managers at the SADC Plant Genetic Resources Centre
(SPGRC) follow the recommendations published by Bioversity International (formerly IPGRI) and other Advanced Research
Centres during a regeneration process. This paper gives an overview of practical considerations and procedures carried out at SPGRC
while regenerating germplasm of orthodox seed crops.

Key words: Genetic integrity, genetic resources, regeneration, seed viability, SPGRC

Introduction characters, with the population then appearing to be more

The Southern Africa Development Community (SADC)
comprises 15 countries with a wide range of agro-climatic
conditions, rich in biodiversity and known for
domestication of several cultivated species. Before the
1980s, there were no formal genebanks for long-term
conservation of genetic resources in the region. The
utilization of modern varieties and agricultural practices
were seen then to be eroding most of this wealth of agro-
biodiversity in the field. SPGRC was established in Lusaka
Zambia in 1989 to hold a base (long term) collection and
facilitate the establishment of National Plant Genetic
Resources Centres (NPGRCs) which hold the active (short
term) germplasm collections in their countries.
A critical objective of a seed conservation programme
is to preserve the genes with the same frequency as found
when they were collected in the field. Seed banks provide
the most practical method for preserving large amounts of
genetic material within a small space. Nonetheless, seeds
in genebanks eventually deteriorate whatever the
conservation conditions. Low temperature and optimizing
seed moisture content can delay seed deterioration, but,
when seed viability decrease to below 85%, regeneration
is recommended. Although the process of regenerating
genetic resources in genebanks may appear routine and a
necessary operation to many scientists, it may lead to
losses of valuable material when conducted without
sufficient attention, to the genetic quality of the accession
thus leading to genetic erosion. Burton 1975 for example
made a comparison of the last generation with the first in
six different germplasm accessions of pearl millet and
showed that advancing germplasm of the crop narrowed
phenotypic variability, lost genes and obscured some
uniform. Therefore, along with the need for seed
regeneration, the process has to be managed in a manner
that the integrity of accessions is maintained as far as
possible. Genebank managers at the SPGRC closely follow
the recommendations published by Bioversity
International and other Advanced Research Centres in
order to maintain the genetic composition of an accession
during a regeneration process. This paper gives an
overview of practical considerations and procedures
carried out at SPGRC while regenerating germplasm of
some orthodox seed crops.
Regeneration procedures
Seed sources for regeneration. National genebanks hold
active collections in their genebanks while SPGRC holds
the region’s base collection. The active collections are
used for evaluation, monitoring, utilization and exchange
purposes. Samples for regeneration are taken from active
collection when seed viability or the amount of seed in
the active collection is depleted. The base collection at
SPGRC is maintained under the most appropriate
conditions for long storage as a back-up in case the
accessions are lost in the active collection. However, no
matter how good the conditions of storage are, the seed
will undergo aging process. Therefore, if viability of seed
in base collection drops below the threshold recommended
by Bioversity, the seed accession will be taken from the
base collection for regeneration.
Criteria for scheduling regeneration of accessions from
base collection. The initial seed germination percentages
for all accessions held in the base collection are known
290 L. LEROTHOLI QHOBELA & W.Y.F. MARANDU
from a register of the internally developed SPGRC
Documentation and Information System (SDIS) database
that is updated each time a sample of any accession is
entered into the collection. Seed accessions in the
genebank are monitored at intervals of five years for
germination percentage and quantity of seed. Accessions
that have low viability and those with insufficient seed
quantities are marked for regeneration and the list is passed
to Regeneration and Multiplication section which will
determine the number of accessions to be regenerated
based on the availability of resources (financial and
personnel) and the type of accessions (wild, weedy,
cultivated) before regeneration is undertaken. First priority
is given to accessions with low seed germination
percentage and insufficient seed quantity; followed by
accessions with low seed germination percentage with
sufficient seed quantity and least prioritized are seed
accessions with high germination percentage with
insufficient seed quantity.
Field practices. Regeneration practices aim to produce
sufficient seed with genetic composition as close to the
original sample as possible; but experience shows that it
is not always possible to produce sufficient seed in one
regeneration cycle from small seed sample sizes commonly
received from collection missions. When the seed sample
size is sufficient the appropriate size of a plot for production
of adequate quantities is chosen. For most field crops the
normal size of a plot is five meters long with 0.80 meters
between rows. The number of rows will vary with the sample
size. On the whole, seed sowing follows the agronomic
requirements of the crop. Proper land preparation, plot
markings in the field and sowing also depend on the crop.
Sowing is done in December and early January and seed
is sown in ridges to encourage drainage. Both basal and
top dressing fertilizers are recommended and
supplementary irrigation is applied during dry spells.
Weeding is done manually and chemicals are used as a
last resort to control pests and diseases. Field rotations
are considered on routine basis to minimize accumulation
and spread of pests and diseases. Strict isolation
procedures to avoid adulteration by foreign pollen (Rao,
et al., 2006) are followed for all cross pollinating
germplasm. Isolation is mainly achieved by controlled
pollination using waxy pollination bags used for collection
of pollen and isolating individual plants or flowers.
Appropriate cost effective mating system such as selfing
as many plants as possible or sibbing is normally used. In
a few cases individual accessions are isolated by distance
or by time. In a number of cases multiple cycles of
regenerations are required to achieve the target amount
of seed for long term storage. Since each multiplication
cycle may affect the genetic composition of the original
accession by altering the frequencies from one generation
to another or by the loss of alleles a target minimum number
of plants to be grown for regeneration is normally set. For
example for pearl millet, at least a minimum of 200 plants
are required from each regeneration cycle while 150 plants
are grown for sorghum accessions 150 plants are grown
out for maize accessions to maintain genetic integrity and
get sufficient seed quantities.
Seed storage facilities and seed longevity. The growing
conditions, post harvesting handling, drying, packaging
and storage conditions determine seed longevity. To
achieve high initial seed viability, infestation of seed by
pest and diseases is minimized. However, some germplasm
accessions are more vulnerable than others and problems
such as diseases and pest attacks are sometimes
encountered. If the problem is experienced for the first
time and appears to have a substantial effect on the plants,
where possible chemicals are applied to control the pests
or diseases otherwise application of chemicals is not
encouraged during seed regeneration activities in the field.
Attention is also paid to harvesting each accession at the
right time, giving priority to shattering types. Post-
harvesting handling activities such as threshing are carried
out with great care before the pre-storage activities begin.
The genebank uses the dehumidification method of drying
germplasm with drying temperature of 150C and relative
humidity of 15% to dry seed to between 5-3% of moisture
content achieved over a period of 2 to 4 weeks for many
crop species in a walk-in drier. The method minimizes
damage to the seed even to seed with high initial moisture
content. When seed has dried to required low moisture
content, it is placed in airtight bottle for conservation and
in five aluminum foil packets each used to monitor viability
status of the accession on intervals of five years or more.
The sealing operation is carried out with care and as fast
as possible to ensure seed does not re-absorb moisture
after drying and before it is stored at -20oC in the genebank.
To store adequate seed in a bottle and ensure sufficient
seed is available for monitoring viability over a period of
30 years, at least 2500 seeds from self pollinated species
and 4000 from cross pollinated species are required. The
genebank uses chest freezers to conserve the base
materials as appropriate technology for the region.
Discussion
Constraints. Regeneration is a costly difficult operation
that requires special skills. Two main factors are considered
at SPGRC during regeneration of germplasm: the quantity
of seed to be produced and preserving the genetic integrity
of the accession. To preserve the genetic integrity of the
accession during seed regeneration, crop specific or
sometimes accession specific procedures are applied.
Theoretical knowledge on the factors that affect genetic
integrity of germplasm under regeneration is very
important. Regeneration guidelines have been developed
for a number of crops. However, the majority of accessions
handled by SPGRC are landraces consisting of mixtures
of populations, therefore, harvesting time and procedures
may not be the best for all the plants which may have an
effect on the longevity of the seeds in storage and
probably unwanted natural selection. This is a major
challenge that must be kept in the minds of staff all the
time to avoid accidental systematic selection. In addition,
 Seed regeneration practices
wild species are particularly difficult to regenerate
optimally. Brown (1997) emphasizes that optimum
regeneration of wild species requires a clear definition of
objectives and priorities among accessions, monitoring
of mating system and genetic structure, maintenance of
accession purity and associated passport data, and
biologically realistic and flexible guidelines for sample size.
Most wild, weedy, and some cultivated species have not
yet been regenerated at SPGRC partly because of this
problem. If an accession cannot be restored from active
collection, the national genebank is normally asked to make
arrangements for regeneration closest to a site where the
sample was adapted and collected, and usually, specialist
scientists who are familiar with the biology of wild, weedy
and other difficult species in the country of origin are
consulted. A site at SPGRC is used to regenerate accessions
with majority being from major cereals crop species
including some maize, sorghum and pearl millets which
grow in a wide range of ecological conditions. Data from
the SPGRC Documentation and Information System (SDIS)
database is used to determine on a case by case basis,
whether an accession can be regenerated at SPGRC fields.
Other constraints include incidences of wild animals
selecting out specific material during growing season
necessitating hiring people to scare them; power shortages
resulting in heavy dependence of the standby generator.
These constraints increase the cost of regeneration
considerably.
Research needs. Considering the number of crops and
the type of materials maintained by the SPGRC, there is
need for studies in the regeneration of mixed populations,
wild relatives of crops and some of lesser known crops.
Paying attention to methods and tools such as use of
controlled pollinations, planting an appropriate number
and making balanced samples from the individuals
contributing seed to the next generation is important for
efficiency and quality of samples put in storage. Dormancy
breaking protocols for some wild, weedy and difficult
291
species including less known crop species need to be
developed. These kinds of studies can be coordinated in
the region as well as outside the region so as to get the
best input by different experts.
Conclusions
Regeneration of genebank accessions by SPGRC heavily
depends on the network linkage with NPGRCs which
provide appropriate sites for materials collected from their
countries and also expertise. By carefully sifting through
the documentation system it has been possible to narrow
down to crops and accessions that can be regenerated at
the SPGRC site allowing NPGRCs to delegate some of the
regeneration load whenever appropriate. A large number
of the materials under conservation are heterogeneous
landraces or wild crop relatives which are not well studied.
These materials therefore pose a major challenge to
scientists since they will soon need regeneration. It is,
therefore, necessary to initiate coordinated studies in the
region as well as outside the region so as to get the best
input by different experts.
Acknowledgement
This paper was prepared with permission of the Head of
SPGRC which is greatly appreciated.
References
Brown, A. H. D., Brubaker, C.L. & Grace, J.P. 1997.
Regeneration of Germplasm Samples: Wild versus
Cultivated Plant Species. Crop Science 37, 7-13
Burton, G..W. 1975. Gene loss in Pearl millet germplasm
pools. Crop Science 16, 251-255.
Rao, N.K., Hanson, J., Dullo, M.E., Ghosh, K., Nowell, D.
& Larinde, M. 2006. Manual of Seed Handling in
Genebanks. Handbook for Genebanks No.8. Bioversity
International Rome, Italy.