[Show abstract][Hide abstract] ABSTRACT: Tef (Eragrostis tef) is a major cereal crop in Ethiopia. Lodging is the primary constraint to increasing productivity in this allotetraploid species, accounting for losses of ~15-45% in yield each year. As a first step toward identifying semi-dwarf varieties that might have improved lodging resistance, an ~6X fosmid library was constructed and used to identify both homoeologues of the dw3 semi-dwarfing gene of Sorghum bicolor. An EMS mutagenized population, consisting of ~21,210 tef plants, was planted and leaf materials were collected into 23 superpools. Two dwarfing candidate genes, homoeologues of dw3 of sorghum and rht1 of wheat, were sequenced directly from each superpool with 454 technology, and 120 candidate mutations were identified. Out of ten candidates tested, six independent mutations were validated by Sanger sequencing, including two predicted detrimental mutations in both dw3 homoeologues with a potential to improve lodging resistance in tef through further breeding. This study demonstrates that high throughput sequencing can identify potentially valuable mutations in under-studied plant species like tef, and has provided mutant lines that can now be combined and tested in breeding programs for improved lodging resistance.
[Show abstract][Hide abstract] ABSTRACT: Tef is one of the staple cereal crops in Ethiopia. To evaluate genetic diversity of tef and its relatives, 47 accessions of
tef, three accessions of E. pilosa, and six accessions of E. curvulawere analyzed using random amplified polymorphic DNA (RAPD) markers. The level of polymorphism among the wild species was
extremely high, while low polymorphism was detected among tef accessions. All cultivars and wild species under study could
be distinguished with the help of different primers, thereby indicating the potential of RAPD in the genetic fingerprinting
of tef. Accessions from E. curvula and E. pilosa can be differentiated by a single selected primer. In spite of low polymorphism within tef, accessions under study could
be distinguished by a combination of selected primers. Cluster analysis indicated that tef is a very closely related species
to E. pilosa with 45%similarity, supporting the hypothesis that tef originated from E. pilosa based on morphological data. Given that RAPD are relatively quick, simple to use, and are not subjected to environmental
influences, they provide a valuable new approach for the genetic fingerprinting and study of genetic diversity in tef.
[Show abstract][Hide abstract] ABSTRACT: The development of single nucleotide polymorphism (SNP) markers in maize offers the opportunity to utilize DNA markers in
many new areas of population genetics, gene discovery, plant breeding and germplasm identification. However, the steps from
sequencing and SNP discovery to SNP marker design and validation are lengthy and expensive. Access to a set of validated SNP
markers is a significant advantage to maize researchers who wish to apply SNPs in scientific inquiry. We mined 1,088 loci
sequenced across 60 public inbreds that have been used in maize breeding in North America and Europe. We then selected 640
SNPs using generalized marker design criteria that enable utilization with several SNP chemistries. While SNPs were found
on average every 43 bases in 1,088 maize gene sequences, SNPs that were amenable to marker design were found on average every
623 bases; representing only 7% of the total SNPs discovered. We also describe the development of a 768 marker multiplex assay
for use on the Illumina® BeadArray™ platform. SNP markers were mapped on the IBM2 intermated B73×Mo17 high resolution genetic map using either the
IBM2 segregating population, or segregation in multiple parent-progeny triplets. A high degree of colinearity was found with
the genetic nested association map. For each SNP presented we give information on map location, polymorphism rates in different
heterotic groups and performance on the Illumina® platform.
[Show abstract][Hide abstract] ABSTRACT: Tef is an important cereal crop in Ethiopia. This study was conducted to investigate (1) genetic diversity within and among three Eragrostis species (E. tef, E. pilosa and E. curvula), and (2) the relationship between E. tef, E. pilosa and E. curvula. A total of 529 AFLP markers were obtained, out of which 58% (368) were polymorphic, using 10 primer, combinations. The three species were separated distinctly using amplified fragment length polymorphism (AFLP), However, diversity revealed at the morphological trait level was not commensurate with that observed for AFLP. This was due to the small number of morphological traits available and their interaction with the environment. Within tef, ‘Rubicunda’ and DZ-01-1093 were found to be distantly related to the rest of the tef accessions. The diversity within species was such that E. pilosa was the most diverse followed by E. curvula and E. tef. Moreover, E. pilosa was more closely related to E. tef than E. curvula. Therefore, further study is needed of E. pilosa accessions and of ‘Rubicunda’ and DZ-01-1093 in a crossing programme to generate a population for selection and/or genetic mapping. A total of 19 cultivars or accessions had one or more unique fragments using one or more AFLP primers, indicating the potential of the technology in fingerprinting tef in a breeding or seed multiplication programme.
[Show abstract][Hide abstract] ABSTRACT: In the fully sequenced Arabidopsis (Arabidopsis thaliana) genome, many gene models are annotated as "hypothetical protein," whose gene structures are predicted solely by computer algorithms with no support from either expressed sequence matches from Arabidopsis, or nucleic acid or protein homologs from other species. In order to confirm their existence and predicted gene structures, a high-throughput method of rapid amplification of cDNA ends (RACE) was used to obtain their cDNA sequences from 11 cDNA populations. Primers from all of the 797 hypothetical genes on chromosome 2 were designed, and, through 5' and 3' RACE, clones from 506 genes were sequenced and cDNA sequences from 399 target genes were recovered. The cDNA sequences were obtained by assembling their 5' and 3' RACE polymerase chain reaction products. These sequences revealed that (1) the structures of 151 hypothetical genes were different from their predictions; (2) 116 hypothetical genes had alternatively spliced transcripts and 187 genes displayed polyadenylation sites; and (3) there were transcripts arising from both strands, from the strand opposite to that of the prediction and possible dicistronic transcripts. Promoters from five randomly chosen hypothetical genes (At2g02540, At2g31270, At2g33640, At2g35550, and At2g36340) were cloned into report constructs, and their expressions are tissue or development stage specific. Our results indicate at least 50% of hypothetical genes on chromosome 2 are expressed in the cDNA populations with about 38% of the gene structures differing from their predictions. Thus, by using this targeted approach, high-throughput RACE, we revealed numerous transcripts including many uncharacterized variants from these hypothetical genes.
[Show abstract][Hide abstract] ABSTRACT: Through comparative studies of the model organism Arabidopsis thaliana and its close relative Brassica oleracea, we have identified conserved regions that represent potentially functional sequences overlooked by previous Arabidopsis genome annotation methods. A total of 454,274 whole genome shotgun sequences covering 283 Mb (0.44 x) of the estimated 650 Mb Brassica genome were searched against the Arabidopsis genome, and conserved Arabidopsis genome sequences (CAGSs) were identified. Of these 229,735 conserved regions, 167,357 fell within or intersected existing gene models, while 60,378 were located in previously unannotated regions. After removal of sequences matching known proteins, CAGSs that were close to one another were chained together as potentially comprising portions of the same functional unit. This resulted in 27,347 chains of which 15,686 were sufficiently distant from existing gene annotations to be considered a novel conserved unit. Of 192 conserved regions examined, 58 were found to be expressed in our cDNA populations. Rapid amplification of cDNA ends (RACE) was used to obtain potentially full-length transcripts from these 58 regions. The resulting sequences led to the creation of 21 gene models at 17 new Arabidopsis loci and the addition of splice variants or updates to another 19 gene structures. In addition, CAGSs overlapping already annotated genes in Arabidopsis can provide guidance for manual improvement of existing gene models. Published genome-wide expression data based on whole genome tiling arrays and massively parallel signature sequencing were overlaid on the Brassica-Arabidopsis conserved sequences, and 1399 regions of intersection were identified. Collectively our results and these data sets suggest that several thousand new Arabidopsis genes remain to be identified and annotated.
[Show abstract][Hide abstract] ABSTRACT: Osmotic adjustment (OA) and deep roots were shown to be important drought resistance mechanism in many crop plants. In this
study, geno types systematically selected from an Ethiopian endemic tef [Eragrostis tef (Zucc) Trotter] germplasm pool were evaluated for osmotic adjustment and root depth in greenhouse in several experiments.
The association of these traits with other plant characters was also studied. Osmotic adjustment was investigated in two experiments.
Experiment 1 was conducted using nine genotypes for two seasons (spring and fall 1996), and experiment 2 was undertaken in
the spring of 1997 with 45 genotypes. In experiment 1, there were significant genotype effects on OA. Though there was also
significant genotype by season interaction for OA, some of the extreme lines gave consistently high (Ada and DZ-01-99) and
low (DZ-01-354 and Trotteriana) OA values across seasons. There was a significant variation among genotypes for OA in experiment
2. Osmotic adjustment was not associated with the altitude of the region of origin of the particular tef genotype. In both
experiments, OA was significantly correlated across tef genotypes with delayed wilting and the maintenance of higher relative
water content (RWC) under conditions of soil moisture stress. Three experiments were conducted to evaluate the phenotypic
diversity for root depth. In all experiments tef genotypes differed significantly for root depth as measured at flowering
time. The late maturing genotypes tended to have greater root depth as compared to early maturing genotypes. Root depth was
not associated with the altitude of the region of origin of the tested genotypes. There was no association between root depth
and OA across genotypes.
[Show abstract][Hide abstract] ABSTRACT: Tef [Eragrostis
tef (Zucc)Trotter] is one of the most important cereal crops in Ethiopia. It is an allotetraploid species with a genome size
of 720 Mbp. In this paper we report results of genetic linkage-map construction for E. tef using tef and heterologous cDNA probes for the first time. One hundred and sixteen F8 recombinant inbred lines (RILs) from the cross E. tef cv Kaye Murri×Eragrostis pilosa (accession30–5) were used for mapping. Parental lines were digested with nine restriction enzymes and screened using 159 tef
cDNA and 162 heterologous probes including the grass genome anchor probes. The polymorphism level between parental lines was
66.9%. One hundred and thirty nine polymorphic probes were hybridized against 116 RILs. Both the tef and the heterologous
probes hybridized well against tef genomic DNA. The linkage map defined 1,489 cM of the tef genome comprising 149 marker loci
distributed among 20 linkage groups. The average interval between markers was 9.99 cM. A fraction (14.8%) of the markers deviated
significantly from the expected segregation. Such a genetic linkage map is useful for tagging economically useful genes in
tef because a wide range of agronomically important traits is segregating within this population. This would enable the use
of a marker assisted breeding strategy which, in turn, will enhance breeding efficiency. Alignment of the tef RFLP map with
the rice RFLP map indicates that a number of syntenic chromosomal fragments exist between tef and rice in which the gene order
was for the most part collinear. The comparative mapping information should enable tef scientists to take advantage of whatever
genetic progress is made on the cereal model species rice.
No preview · Article · Apr 2001 · Theoretical and Applied Genetics
[Show abstract][Hide abstract] ABSTRACT: A total of 3600 entries of tef [Eragrostis tef (Zucc.) Trotter] germplasm collections representing 36 populations from six regions of Ethiopia were evaluated for phenotypic diversity in six qualitative and phenologic characters. High values of Shannon–Weaver diversity index (
) were recorded for most characters in each population. Monomorphism was high for anther color (purple) and panicle form (loose). Larger variances for
were obtained among regions than among populations within a region from hierarchical analysis of variance. Significant (P<0.05) to highly significant (P<0.01) t-values were observed among
values of some regions for seed color, panicle form and days to maturity. A substantial level of variance (P<0.05) was obtained for populations within altitudinal zones only for days to maturity. Altitudinal zones exhibited similar levels of diversity across characters for these collections. Generally, considerable variations important for tef improvement work have been observed and regions with highest diversity for some traits have been suggested for future in situ or ex situ germplasm conservation works. Although similar mean diversity indices were obtained among regions, they were non overlapping.
No preview · Article · Jan 2000 · Genetic Resources and Crop Evolution
[Show abstract][Hide abstract] ABSTRACT: A genetic linkage map of tef was constructed with amplified fragment length polymorphism (AFLP) markers using F(5) recombinant inbred lines (RILs) derived by single seed descent from the intraspecific cross of 'Kaye Murri'×'Fesho'. A total of 192 EcoRI/MseI primer combinations were screened for parental polymorphism. Around three polymorphic fragments per primer combination were detected, indicating a low polymorphism level in tef. Fifty primer combinations were selected to assay the mapping population, and 226 loci segregated among 85 F(5) RILs. Most AFLP loci behaved as dominant markers (presence or absence of a band), but about 15% of the loci were codominant. Significant deviations from the expected Mendelian segregation ratio were observed for 26 loci. The genetic linkage map comprised 211 markers assembled into 25 linkage groups and covered 2,149 cM of genome. AFLP is an efficient marker system for mapping plant species with low polymorphism such as tef. This is the first genetic linkage map constructed for tef. It will facilitate the mapping of genes controlling agronomically important traits and cultivar improvement in tef.
No preview · Article · Aug 1999 · Theoretical and Applied Genetics
[Show abstract][Hide abstract] ABSTRACT: Tef, [Eragrostis tef(Zucc) Trotter, is a major cereal crop in Ethiopia. The objective of this study was to investigate the genetic relationships among tef accessions by the amplified fragment length polymorphism (AFLP) technique. Forty-seven accessions of tef and one accession each of E. pilosa and E. curvula were analyzed. A total of 316 polymorphic DNA fragments across the three Eragrostis species were identified. The level of polymorphism within tef accessions was low (18%). However, with four or more primer pairs and a cluster analyses procedure, all the accessions included in the study were differentiated from each other. The three species studied fell in to three distinct clusters. Within tef, two major groups can be differentiated at 89% similarity. Because AFLP is reliable and enables assessment of potentially unlimited numbers of genetic loci, it permits greater insights into the genetic diversity of tef than had been possible with other available markers. Despite the low polymorphism level in tef, the availability of a large number of primer combinations and the high speed of the assay make AFLP an efficient marker technology for genetic diversity study of tef.
[Show abstract][Hide abstract] ABSTRACT: Three hundred twenty germplasm lines of the major Ethiopian cereal, tef, [ Eragrostis tef (Zucc.) Trotter], were evaluated
for 20 morphological, phenological and agronomic characters in two-replicated randomized complete blocks at Debre Zeit and
Melkassa Agricultural Research Centers in Ethiopia during the 1995 main season. The objectives were to assess the diversity
of the lines, and to estimate the broad sense heritability (H) and genetic advance (GA) of the various characters. The mean
squares of genotypes were highly significant (p ≤ 0.001) for all the traits. The phenotypic and genotypic coefficients of
variation ranged in that order from about 6–40% and 3–23% for days to maturity and grain yield/plant, and days to maturity
and number of spikelets/main shoot panicle, respectively. The cluster analysis grouped the genotypes into 14 major complexes
consisting of one to 183 lines. Of the 19 principal components involved in explaining the entire variation among the genotypes
the first five which had eigenvalues of more than one explained about 73.8%. The first principal component which accounted
for about 34% of the total variance was due chiefly to plant height, culm and panicle length, diameters of the two basal culm
internodes, main shoot panicle mass and grain yield, number of main shoot panicle branches and spikelets, and days to panicle
emergence and maturity. Estimates of H varied from about 22% for length of the lowest main shoot stem internode to 74% for
number of main shoot panicle branches. Values of GA (expected from selection of the superior 5% of the lines and expressed
relative to the means) ranged from about 3% for days to maturity to 36% for number of spikelets/main shoot panicle. Overall,
the study indicated the existence of trait diversity in tef germplasm and this can be exploited in the genetic improvement
of the crop through hybridization and selection.
[Show abstract][Hide abstract] ABSTRACT: Tef [Eragrostis tef (Zucc.) Trotter] is the most important cereal crop in Ethiopia. An experiment was conducted to investigate genetic diversity among four cultivars of tef and 14 accessions of Eragrostis pilosa using radiolabelled and silver stained amplified fragment length polymorphism. Morphological traits were also evaluated. A total of 897 markers were obtained out of which 395 were polymorphic using 11 primer combinations. Cluster analysis revealed accessions of E. pilosa which are distantly related and others closely related to tef. Our previous experience also indicates that E. pilosa is crossable with tef. Those accessions distantly related to tef could be used in a crossing program to generate a population for selection and/or genetic mapping. Such genetic mapping populations will form an important entry point towards the molecular genetic dissection of the plant genus, Eragrostis, especially in the context of comparative mapping. Knowledge gained from such study, apart from tef improvement, will also be useful for many forage and turf grass species where little molecular genetic information is available. Nine cultivars or accessions had one or more unique fragments using one or more AFLP primers indicating the potential of the technology in fingerprinting tef in a breeding or seed multiplication program. The results also showed that clusters obtained using silver staining and gamma 33P-ATP labeling were similar, suggesting that silver staining could be used as an alternative to radiolabeling at least in genetic diversity analysis. Significant genetic variation was obtained for morphological traits. Of particular interest to tef breeding was short plant stature in E. pilosa which could be transferred to tef to minimize the problem of lodging. Diversity revealed at the morphological trait level was not commensurate with that observed for AFLP. This was due to the small number of available morphological traits and their interaction with the environment.
[Show abstract][Hide abstract] ABSTRACT: Flow cytometric analysis of nuclear DNA content was performed by using nuclei isolated from young leaf tissue of tef (Eragrostis tef). The method was very useful for rapid screening of ploidy levels in cultivars and lines of tef representing the phenotypic variability of this species in Ethiopia. The results of the analysis showed that all cultivars were tetraploid. Flow cytometry was also used to determine nuclear DNA content in absolute units (genome size) in four tef cultivars. Nuclei isolated from tomato (Lycopersicon esculentum, 2C=1.96 pg) were used as an internal reference standard. The 2C DNA content of individual tef cultivars ranged from 1.48 to 1.52 pg (1C genome size: 714 Mbp-733 Mbp), the differences among them being statistically nonsignificant. The fact that the nuclear genome of tef is only about 50% larger than that of rice should make it amenable for analysis and mapping at the molecular level.