1. Outcrossing rates of wild and weedy sorghum (Sorghum bicolor)
in Ethiopia: implications for crop-wild gene flow
A. Adugna1 , A.A. Snow,2 and P. M. Sweeney2
1 Addis Ababa University, Department of Biology, Addis Ababa, Ethiopia, 2Dept. of EEOB, Ohio State Univ. Columbus, OH
ABSTRACT
MATERIALS and METHODS
Gene flow via pollen allows crop genes, including transgenes, to introgress into sexually compatible wild or weedy populations that occur nearby. Because transgenic sorghum (Sorghum bicolor L.) is being developed for Africa, we are investigating the potential for transgenes to spread to conspecific wild/weedy sorghum populations in Ethiopia, the center of origin for cultivated sorghum. Our previous studies showed that wild and cultivated sorghum often co-occur and flower synchronously in Ethiopia. Furthermore, crop-wild hybrids from artificial crosses involving accessions are fertile and vigorous. In the current study, we investigated the extent of outcrossing in six wild/weedy sorghum populations collected from a total of five geographical regions in Ethiopia (Ghibe, Hararghe, Pawe, Tigray, and Wello) at elevations ranging from m. We collected DNA samples from 20 maternal plants per population and 8 progeny plants from each of the twenty maternal genotypes. Data from five polymorphic microsatellite markers were used to estimate outcrossing rates based on Ritland’s multilocus mating system program. The average multilocus outcrossing rate (tm) was 50.5%, with a range of 31-65% among populations, which is higher than reported earlier for cultivated sorghum. Previous studies of sorghum landraces in Cameroon, Morocco, and Burkina-Faso also report variable outcrossing rates, ranging from ~5-40%. The highest outcrossing we observed (65%) was recorded in a weedy population that was intermixed with an improved crop variety in Abuare (Wello) region. Biparental inbreeding was evident in all populations and averaged 23.5% (range = 10-33%). The high outcrossing rates of wild/weedy sorghum populations in Ethiopia indicate a high potential for crop-to-wild gene flow. Effective risk management strategies may be needed if the introgression of transgenes or other crop genes from improved cultivars into wild or weedy populations is deemed to be undesirable.
Collections
Five Regions (7 populations) in Ethiopia (Figure 1)
Ghibe (1population)
Haraghe (1 population)
Pawe (1 population)
Tigray (2 populations: Humera & HagereSelam)
Wello (2 populations: Kobo & Abuare)
Sorghum bicolor ssp. arrundinacae
Sorghum bicolor ssp. drummondii
DNA was collected from 20 Individuals per population
using Whatman FTA or chromatography cards3
Seed heads were collected from the same 140 plants
Germination and Seedling Sampling
8 progeny per each maternal parent were germinated
Samples were taken by squashing 3-8 week old seedling
leaf issue onto Whatman FTA cards or chromatograpy paper
DNA Extraction, PCR Amplification and Analysis
Modified Whatman FTA CardExtraction3
5 Microsateillite Loci
QIAGEN universal mutiplex cycling protocol
ABI 3100 Genetic Analyzer
GeneMapper 3.7 software
Figure 1. Map of Ethiopia showing locations where wild and weedy sorghum populations were sampled
Abuare
Kobo
Ghibe
Hararghe
Hagere-Selam
Pawe
Humera
INTRODUCTION
Sorghum is an important grain world wide. It is drought resistant and is used for food, feed fiber and fuel, in Ethiopia, it’s center of origin where wild and weedy population still exist.. Thus, information about crop-to-wild gene flow in Ethiopia is needed for:
-In situ conservation of wild germplasm
-Risk assessment of novel transgenes
Steps in assessing crop-wild gene flow potential:
-Geographic surveys in center of origin
-Determination of the potential for outcrossing between crop/wild sorghum
-Determination of outcrossing rate of current wild/weedy sorghum populations
-Determine relative fitness of crop-wild hybrids
Previous findings:
-Wild/weedy sorghums were present and intermixed with crop at many sites in Ethiopia.
-Flowering times of wild/weedy sorghums and crop overlapped.
-Sorghum cultivars readily pollinate wild sorghum accessions
-Hybrids between crop and wild accessions are as fit as parental accessions
Objectives
-Estimate the in situ rate of outcrossing of wild and weedy sorghum populations in Ethiopia
Outcrossing Estimates
Based on mixed mating model4
Multi-locus mating system program: MLTR5
RESULTS
Multilocus outcrossing rate ranged from 19.0% (Kobo) to 65.4% (Abuare)
Highest outcrossing rate recorded in Abuare where weedy sorghum was intermixed with improved cultivar.
Average multilocus outcrossing rate (excluding Kobo where one primer pair failed to amplify) was 50.5%
Table. 1 Estimates of outcrossing rate in wild and weedy sorghum populations in Ethiopia
** average doesn’t include Kobo families
Outcross
Estimate
Ghibe
Harangue
Pawe
Humera
/Donkey
Hagere-Selam
Abuare
Kobo
Average**
Multi-locus
0.569
(0.071)
0.479
(0.070)
0.453
(0.054)
0.331
(0.065)
0.609
(0.058)
0.654
0.190
(0.072)
0.505
(0.026)
Single Locus-
0.242
(0.037)
0.231
(0.041)
0.265
(0.033)
0.230
0.343
(0.062)
0.376
(0.045)
0.138
(0.067)
0.205
(0.013)
Multi-locus –Single Locus
0.328
(0.040)
0.248
0.187
(0.028)
0.101
(0.022)
0.266
(0.039)
0.278
(0.032)
0.051
(0.018)
0.300
(0.016)
References
CONCLUSIONS
1. Tesso, T., I Kapran, C. Grenier, A. Snow, P. Sweeney, J. Pederson, D. Marx, G. Bothman, N
And G Ejeta The potential for crop-to-wild gene flow in sorghum in Ethiopia and Niger: a geographic survey. Crop Sci 48:
2. Snow, Allison A., Patricia Sweeney, Cecile Grenier, Geibisa Ejeta, Tesfay Tesso, Issofrou Kapran, Gurling Bothma, and Jeffery F. Pedersen Lifetime fecundity of F1 crop-wild sorghum hybrids: Implications for gene flow from transgenic sorghum in Africa.  Tenth International Symposium on the Biosafety of Genetically Modified Organisms
3. Adugna, A., A. Snow and P. Sweeney. Optimization of a high throughput, cost effective, and all-stage DNA extraction protocol for sorghum (Sorghum bicolor). Journal of Agricultural Science and Technology ( accepted).
4. Brown, A. and R. Allard The estimation of Wright’s fixation index from genotypic frequencies. Genetica 41:
5. Ritland, K
Relatively high average outcrossing rate suggests the potential for crop to wild gene flow in sorghum.
Outcrossing rates vary among the wild sorghums populations, suggesting that populations may vary in their potential to accept pollen from nearby crops.
High outcrossing rate in Abuare, where a wild sorghum population was intermixed with improved cultivar, indicate a high risk of gene flow from sorghum crop in this area.
Effective risk management strategies are needed to monitor and contain gene flow from future GM sorghum to non-GM wild and weedy sorghum populations in Ethiopia.