A REAPPRAISAL OF THE SUBGENUS GLYCINE

The genus Glycine Willd. is divided into three subgenera, Glycine Willd., Soja (Moench) F. J. Herm., and Bracteata Verdc. Six species are currently recognized in the subgenus Glycine: G. canescens F. J. Herm., G. clandestina Willd., G. falcata Benth., G. latrobeana (Meissn.) Benth., G. tabacina (Labill.) Benth., and G. tomentella Hayata. Distribution of the subgenus extends from south China to Tasmania and includes several Pacific islands. A collection of these species was examined cytologically and morphologically in an attempt to evaluate existing variability between and within taxa. Chromosome counts confirmed G. canescens, G. clandestina and G. falcata to be diploid with 2n = 40. Both tetraploids (2n = 80) and diploids were found in G. tabacina, the latter restricted to Australia. Glycine tomentella accessions were primarily tetraploid, but several collections from New South Wales, Australia, were found to be aneuploid with 78 chromosomes. One collection was aneuploid at the diploid level with 38 chromosomes. Meiosis appeared normal in the aneuploids with regular bivalent formation. Several accessions previously identified as G. tomentella were diploid. Seed of G. latrobeana was not available for analysis. Numerical techniques in the form of cluster analysis and principal components analysis were applied to morphological data on vegetative and inflorescence characters obtained from each collection. Numerical analysis grouped the accessions essentially according to current species delimitations with some exceptions. Glycine tabacina specimens from Taiwan approached G. clandestina in several characteristics. The diploid G. tomentella specimens formed a separate cluster and appeared morphologically distinct from the remaining taxa.     

Distribution of rDNA loci in the genus Glycine Willd.

The objective of this study was to examine the distribution of rDNA loci in the genus Glycine Willd. by fluorescent in situ hybridization (FISH) using the internal transcribed spacer (ITS) region of nuclear ribosomal DNA as a probe. The hybridized rDNA probe produced two distinct yellow signals on reddish chromosomes representing two NORs in 16 diploid (2n=40) species. Aneudiploid (2n=38) and aneutetraploid (2n=78) Glycine tomentella Hayata also exhibited two rDNA sites. However, the probe hybridized with four chromosomes as evidenced by four signals in two diploid species (Glycine curvata Tind. and Glycine cyrtoloba Tind.) and tetraploid (2n=80) G. tabacina (Labill.) Benth. and G. tomentella. Synthesized amphiploids (2n=80) of Glycine canescens F. J. Herm. (2n=40) and the 40-chromosome G. tomentella also showed four signals. This study demonstrates that the distribution of the rDNA gene in the 16 Glycine species studied is highly conserved and that silence of the rDNA locus may be attributed to amphiplasty during diploidization and speciation. Received: 10 October 2000 / Accepted: 6 December 2000     

The genomic relationships among six wild perennial species of the genus Glycine subgenus Glycine Willd.

Summary Based on meiotic chromosome behavior in intra- and interspecific hybrids, genome symbols were assigned to the following diploid (2n=40) Glycine species: G. canescens = AA; G. clandestina- Intermediate pod (Ip)=A ₁ A ₁; G. clandestina-Short pod (Sp)=BB; G. latifolia = B ₁ B ₁; G. tabacina = B ₂ B ₂; G. cyrtoloba = CC; and G. tomentella = DD. Genome symbol GG was reserved for the soybean, G. max. At metaphaseI, loose chromosome associations were observed in completely sterile interspecific hybrids whose parents differed in their genomes, suggesting some chromosome homologies among species. Although G. clandestina-Sp, G. latifolia and G. tabacina are morphologically distinct species, they differ only by a paracentric inversion. Similar observations were recorded for G. canescens and G. clandestina-Ip. Evidence is presented that demonstrates that G. tabacina (2n=80) and G. tomentella (2n=78, 80) are allotetraploid species complexes. Hybrid weakness, sterility, seedling lethality and seed inviability were found in intra- and interspecific hybrids.This research was supported in part by the Illinois Agricultural Experiment Station and the U.S. Department of Agriculture (Special grant 82-CRSR-2-2007). Travel grants to collect Glycine germplasm were received from the Rockefeller Foundation, the Illinois Soybean Program Operating Board, the National Science Foundation (INT76-14753) and the International Board for Plant Genetic Resources     

Seed coat variation in Glycine Willd. subgenus Glycine (Leguminosae) by SEM

Seed of the genusGlycine Willd. typically exhibits a muriculate appearance resulting from adherence to the true seed coat of the perisperm or inner pod wall layer. Thickened cell walls of the perisperm superimpose a reticulate network on the seed coat, the type of network ranging from alveolate to stellate depending on the shape of the perisperm cells. Tubercles distributed at intervals give the seed its roughened appearance. Seed lacking an attached perisperm appears smooth and shiny. Seed morphology of 62 collections representing the six species of the subgenusGlycine is examined in detail to elucidate inter-and intraspecific variability. Seed perisperm pattern appears to be characteristic for each species, but there are exceptions.Glycine canescens F. J. Herrn. andG. clandestina Willd. seeds possess a reticulate network and tubercles of irregular shape, the perisperm appearing granular inG. clandestina. Seeds ofG. latrobeana (Meissn.) Benth. andG. tabacina (Labill.) Benth. lack a distinct network and have stellate tubercles; the perisperm is granular inG. latrobeana and some plants ofG. tabacina. A few collections ofG. clandestina approachG. tabacina in seed appearance.Glycine tomentella Hayata seeds exhibit a regularly alveolate arrangement, while those ofG. falcata Benth. lack a perisperm layer altogether. Variation in seed coat within a species can usually be linked to differences in chromosome number or some aspect of gross morphology. Diploid collections ofG. tomentella (2n = 40) exhibit recognizable differences in seed morphology compared with tetraploids (2n = 80), coincident with other striking dissimilarities in gross morphology. An incompletely attached perisperm is accompanied by aneuploidy in severalG. tomentella accessions, while other 78 and 38 chromosome aneuploids produce normal seeds.     

Genomic relationships in the genus Glycine (Fabaceae: Phaseoleae): use of a monoclonal antibody to the soybean Bowman-Blrk inhibitor as a genome marker

We investigated the use of a monoclonal antibody (MAb 238) to the soybean Bowman-Birk inhibitor (BBI) to verify and understand the intergenomic relationships among the wild perennial Glycine species. Competitive enzyme linked immunosorbent assay and western blot screening studies revealed that the accessions of B-genome (G. latifolia, G. microphylla, and G. tabacina, 2n = 40) and C-genome (G. curvata and G. cyrtoloba) species did not contain the MAb 238 crossreactive proteins (BBI-nulls). By contrast, all the A-genome (G. argyrea, G. canescens, G. clandestina, and G. latrobeana), E-genome (G. tomentella, 2n = 38), and F-genome (G. falcata) species, G. arenaria (genome unknown), and the polyploid (2n = 78,80) G. tomentella accessions were BBI-positive. The D-genome G. tomentella (2n = 40) and tetraploid G. tabacina (2n = 80) contained both BBI-null and BBI-positive type accessions. Among the recently described species, G. hirticaulis (2n = 40), G. lactovirens, and G. pindanica contained the MAb 238 crossreactive proteins while G. albicans did not. Glycine hirticaulis, G. pindanica, and G. tomentella (2n = 38) displayed highly similar MAb 238 crossreactive isoelectric focusing banding patterns, indicating that they are genomically close to each other. Glycine hirticaulis was found to have both diploid (2n = 40) and tetraploid (2n = 80) cytotypes. We demonstrated that the MAb 238 was specific to the trypsin inhibitor domain of the BBI. The MAb 238 clearly reflected all the previously established relationships in the genus Glycine, validating its use as a genome marker.     

Variation in the DNA Content of Glycine Species

Nuclei were isolated from cotyledons of a range of accessionsfrom 14 species of Glycine. These were stained with ethidiumbromide and the relative fluorescence for each genotype wasmeasured by flow cytometry. The DNA content was estimated bycomparison of relative fluorescence with that from nuclei fromseedling leaves of Allium cepa, whose DNA content has been calculatedpreviously by chemical assay. The 4C amounts for diploid Glycineranged from 3.80 to 6.59 pg. Two groups of diploid species appearedfrom the analysis. The first consisted of species with amountsranging from 3.80 to 5.16 pg and included G. canescens (AA),G. argyrea (A₁ A₁), G. clandestina (A²A²), G. microphylla(BB),G. latifolia (B₁B₁), G. tabacina 2n=40 (B²B²), G. tomentella2n=38 (EE) and 2n=40 (DD), G. max and G. soja (GG), G. arenariaand G. latrobeana. A second group had higher DNA contents rangingfrom 5.27 to 6.59 pg, and consisted of G. curvata, G. cyrtoloba(CC), and G. falcata (FF). The polyploid species, G. tabacina2n=80 (AABB, BBB¹B¹), G. tomentella 2n=78 and 2n=80 (AAEE andDDEE, respectively) contained amounts approximating to the sumsof the respective parental diploid species thought to have givenrise to these allotetraploids. Intraspecific variation was detectedin the DNA content of G. canescens. Within the overall distributionof DNA amounts found in A genome species, each genome containeda range of DNA contents specific to that species. This phenomenonwas also detected amongst B genome species.     

Systematic Status of the Glycine tomentella and G. tabacina Species Complexes (Fabaceae) Based on ITS Sequences of Nuclear Ribosomal DNA

Glycine was reconstructed based on nucleotide sequences of the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA to examine the systematic status of the G. tomentella and G. tabacina species complexes. Rooted at the subgenus Soja (2 species, 7 accessions), parsimony analysis was conducted for 17 species (31 accessions) of the subgenus Glycine, including 9 and 6 populations of G. tomentella s.l. and G. tabacina s.l., respectively. The nrlTS phytogeny indicated polyphyly of G. tomentella s.l. as well as G. tabacina s.l. In the G. tomentella species complex, larger legumes, narrower leaflets, and deflexed indumentum hairs differentiated G. dolichocarpa from G. tomentella s.s. The tetraploid G. dolichocarpa (2n=80) and aneuploid G. tomentella (2n= 38) represented independent lineages from another clade of the remaining diploid (2n=40) and tetraploid species with a DD genome type. Tetraploid G. tabacina (2n=80) was closely related to G. dolichocarpa instead of the diploid G. tabacina (2n=40) with a BB genome type. The nrlTS phytogeny suggested allopolyploidy of G. dolichocarpa and of the tetraploid G. tabacina, both of which possibly share a common parental species with an AA genome type. Their phylogenetic affinity also indicated biased inheritance of the nrDNA ITS and a possible dominant role of the AA genome. Phylogenetically, G. dolichocarpa and allotetraploid G. tabacina should be recognized as distinct species. Received 12 February 2001/ Accepted in revised form 17 August 2001     

Adventitious roots inGlycine subg.Glycine (Leguminosae): Morphological and taxonomic indicators of the B genome

Adventitious roots were observed on 3 wild perennialGlycine species, in short-poddedG. clandestina, G. latifolia and mostG. tabacina (2x, 4x, 6x), while other species lacked adventitious roots. This suggests that the adventitious roots trait is associated with B genome species. Intra- and interspecific F₁ hybrids reveal that adventitious roots apparently are inherited recessively. The presence of adventitious roots or short-poddedG. clandestina, coupled with infertility with longer-poddedG. clandestina, and enzymatic and leaflet morphology differences between the twoG. clandestina subgroups, supports segregation of the short-podded form as a separate taxon.     

A CHLOROPLAST-DNA PHYLOGENY OF THE WILD PERENNIAL RELATIVES OF SOYBEAN (GLYCINE SUBGENUS GLYCINE): CONGRUENCE WITH MORPHOLOGICAL AND CROSSING GROUPS

Hypotheses of evolutionary relationships among the Australian wild perennial relatives of soybean (Glycine subgenus Glycine) are based largely on patterns of meiotic pairing in intra- and interspecific experimental hybrids. This evidence has indicated a number of genome groupings within the subgenus but has not resolved most phylogenetic relationships. Restriction-endonuclease site variation of chloroplast DNA (cpDNA) within the perennial subgenus is reported here, representing a sampling of approximately 3% of the approximately 150-kilobase plastome. Seven hundred twenty-one unique restriction sites were compared within Glycine using 29 restriction endonucleases; 157 sites varied within the genus. Distance and parsimony methods using these data yielded congruent results, recognizing the existence of three major groups within subgenus Glycine: the species-rich and geographically diverse A clade consisting of G. canescens and related taxa; the B clade, which includes the stoloniferous species; and the C group, containing two species with distinctive curved pods. These results are in general agreement with hypotheses based on genome analysis; inconsistencies involve the inclusion of genetically divergent taxa such as G. falcata in well-supported plastome clades comprised of otherwise interfertile species. Such findings are not unexpected if crossing barriers are considered to be unique features of such anomalous species, paralleling their often numerous morphological and cpDNA autapomorphies. Consideration of cpDNA divergence within the three major clades of subgenus Glycine indicates that the rate of plastome evolution is uncoupled from rates of morphological or ecological diversification.     

Intersubgeneric hybridization of soybeans with a wild perennial species,Glycine clandestina Wendl

Summary The exploitation of wild perennial species of subgenus Glycine has been formidable in soybean breeding programs because of extremely poor crossability and an early pod abortion. The combination of gibberellic acid application to hybridized gynoecia and improved seed culture media formulations resulted in a new intersubgeneric hybrid between Glycine max (L.) Merr. (2n=40) and G. clandestina Wendt. (2n=40). Of the 31 immature seeds cultured, 1 regenerated 21 plants through organogenesis while the remaining 30 failed to germinate. All the regenerated plants were similar morphologically, carried expected 2n=40, possessed hybrid isozyme patterns and were completely sterile. Complete absence of chromosome pairing was observed in 40.9% sporocytes. The occurrence of 1 to 6 loosely paired rod bivalents suggests some possibilities of allosyndetic pairing. Hybrid plants set aborted pods after backcrossing to G. max.     

Leaf flavonoids of Glycine subgenus Glycine in relation to systematics

A survey of leaf flavonoids and isoflavonoids in several taxa of the genus Glycne subgenus Glycine was undertaken to see if this would help interpret inter- and intraspecific relationships in the genus. C-Glycosylflavones based on apigenin were found in Glycine tomentelia, G. tabacina and G. falcata. Glycosides of quercetin and kaempferol were also detected in G. tabacina. In the cultivated soybean, G. max, and its wild annual relative, G. soja, only quercetin and kaempferol glycosides have been reported. Interspecific hybrids of Glycine species sometimes show additive flavonoid patterns in F₁ hybrid leaf tissue. All perennial wild species analysed including Glycine canescens and G. latifolia have the isoflavonoids genistin (genesitein 7-O-glucoside), daidzein and coumestrol in the leaves.     

SSR marker and ITS cleaved amplified polymorphic sequence analysis of soybean × Glycine tomentella intersubgeneric derived lines

Wild perennial Glycine species are an invaluable gene resource for the cultivated soybean [Glycine max (L.) Merr., 2n=40]. However, these wild species have been largely unexplored in soybean breeding programs because of their extremely low crossability with soybean and the need to employ in vitro embryo rescue methods to produce F₁ hybrids. The objective of this study was to develop molecular markers to identify gene introgression from G. tomentella, a wild perennial Glycine species, to soybean. A selection of 96 soybean simple sequence repeat (SSR) markers was evaluated for cross-specific amplification and polymorphism in G. tomentella. Thirty-two SSR markers (33%) revealed specific alleles for G. tomentella PI 483218 (2n=78). These SSR markers were further examined with an amphidiploid line (2n=118) and monosomic alien addition lines (MAALs), each with 2n=40 chromosomes from soybean and one from G. tomentella. The results show that the use of SSR markers is a rapid and reliable method to detect G. tomentella chromosomes in MAALs. We also developed a cleaved amplification polymorphism sequence (CAPS) marker according to the sequences of internal transcribed spacer (ITS) regions in soybean and G. tomentella. Four MAALs that carry the ITS (rDNA) locus from G. tomentella were identified. The SSR and ITS-CAPS markers will greatly facilitate the introgression and characterization of gene transfer from G. tomentella to soybean.Communicated by F.J. Muehlbauer     

Wide hybridization between Brazilian soybean cultivars and wild perennial relatives

Employing a different culture strategy, we obtained a greatly improved frequency of embryo rescue in intersubgeneric soybean hybrids. Successful crosses were obtained in 31 different genotype combinations between nine Brazilian soybean lines as the female parents and 12 accessions from Glycine canescens, G. microphylla, G. tabacina and G. tomentella. The hybrid pod retention rate dropped to about 10% during the first 8 days after pollination and stayed largely unchanged up to the 20th day. Immature harvested seeds fell into three size groups: Group 1, smaller than 1.3 mm (mostly empty seed coats); Group 2, 1.9–5.0 mm; Group 3, larger than 5 mm (from selfing). A total of 90 putative hybrid embryos were rescued using a highly enriched B5 medium to nourish the newly dissected embryos. The growing embryos were then placed in a high osmotic, modified B5 medium to induce maturation and dormancy. Schenk and Hildebrandt medium was used to germinate the dormant, partially dehydrated, physiologically mature embryos. Approximately 37% of the rescued embryos developed into plantlets in vitro, and approximately 8% grew into mature plants in the greenhouse. Morphological, cytological and isoenzyme patterns confirmed the hybrid status of all seven mature plants, all of which were generated using G. tomentella G 9943 as the paternal parent. It was observed that all soybean lines crossed with G 9943 were capable of producing mature hybrid plants. There was no correlation between the initial size of Group 2 seeds and plant survival rate. The hybrids were cloned by grafting and treated with colchicine. One of the treated plants displayed chromosome doubling.     

Phylogenetic relationships of the chloroplast genomes in the genus Glycine inferred from four intergenic spacer sequences

The nucleotide sequences of four intergenic spacer regions of chloroplast DNA, atpB-rbcL, trnS-trnG, rps11-rpl36, and rps3-rpl16, were analyzed in the genus Glycine. Phylogenetic analysis based on the sequence data using Neonotonia wightii as the outgroup generated trees supporting the classification of two subgenera, Soja and Glycine, and three plastome groups in the subgenus Glycine. The results were consistent with the presence of diversified chloroplast genomes within tetraploid plants of G. tabacina and G. tomentella, as well as with a close relationship between G. tomentella and G. dolichocarpa that had been suggested based on morphological analyses. Little sequence variation was found in the subgenus Soja, suggesting that G. soja rapidly expanded its distribution in East Asia. The analysis also showed that the differentiation into three plastome groups in the subgenus Glycine occurred in the early stages of its evolution, after the two subgenera diverged.     

Plant regeneration of wildGlycine species from suspension culture-derived protoplasts

Protoplasts isolated from four-week old cell suspension cultures ofGlycine canescens F. J. Herm andG. clandestina Wendl. were cultured in 8P or modified 8P to a multicellular stage. Colonies of 0.5 to 1.0 mm diameter were transferred to solid media for callus growth and regeneration. Callus consisted of friable masses with compact green nodular areas. Organogenesis of both species occurred primarily from the green nodular areas. Shoot buds ofG. clandestina did not mature, but shoots ofG. canescens proliferated on MS medium, with B5 vitamins, 0.33 mgL^–1 each BA, KN, ZN, and 0.15 mgL^–1 NAA. Shoots failed to root after multiple subcultures on four different rooting media.In vitro grafting ofG. canescens scions ontoG. max root stocks allowed plants to be transferred to soil. An overall protoplast division efficiency of 48% was achieved with moderately efficient shoot regeneration inG. canescens. Division efficiencies forG. clandestina were lower (11%). Refinements of this protocol should result in high efficiencies of regeneration which would allowin vitro manipulations of these wild soybean relatives at the single cell level and would make the derivation of somatic hybrid plants possible within the genusGlycine.Abbreviations BA 6-Benzyladenine - KN kinetin - ZN Zeatin - NAA Napthaleneacetic acid - 2,4-D 2–4-dichlorophenoxyacetic acid - PIC Picloram - CH casein hydrolysate - Gln glutamine - Met methionine - MES 2[N-morpholine] ethanesulfonic acid     

Phylogeny of 12 species of genusGlycine Willd. reconstructed with internal transcribed region in nuclear ribosomal DNA

The ITS-Is of 24 accessions belong to 10 species of subgenusGlycine, and 2 species of subgenusSoja of genusGlycine were amplified, cloned and sequenced. According to the homology of the sequences, the phylogeny of the 24 accessions were reconstructed. The reconstructed dendrogram showed that there were some divergent genomic types found in the previously classified species, such asG. tomentella, G. canescens andG. tabacina, and they might be some cryptic species by morphologic analysis. Project supported by the Chinese Academy of Sciences     

Genomic relationships among diploid wild perennial species of the genus Glycine Willd. subgenus Glycine revealed by crossability,meiotic chromosome pairing and seed protein electrophoresis

Summary The nomenclature of species beased on classical taxonomy can be verified from cytogenetic, biochemical and molecular studies. The objective of the study presented here was to provide further information on genomic affinities among species of the genus Glycine Willd. based on crossability, meiotic chromosome pairing of F₁ hybrids and seed-protein profiles. Meiotic chromosome pairing data revealed no genomic similarity between G. microphylla (BB) and G. falcata (FF), nor between G. tomentella (2n = 38; EE) and G. microphylla (BB). Despite morphological similarity between G. cyrtoloba (CC) and G. curvata no F₁ hybrid was obtained, although 748 flowers were pollinated. The seed-protein banding patterns showed G. latrobeana to be closer to the A-genome species than to others. Based on these results we assign genome symbol A₃A₃ to G. latrobeana. Likewise, G. curvata was allotted the designation C₁C₁ because the seed-protein banding patterns of G. curvata and G. cyrtoloba are similar. The genome designations of Glycine species based on cytogenetic investigations may be further extended by results obtained from biochemical and molecular approaches.Research supported in part by the Illinois Agricultural Experiment Station and US Department of Agriculture Competitive Research Grant 88-37231-4100     

Genome relationships between Hordeum vulgare L. and H. bulbosum L.

Interrelationships between H. vulgare (2x=14) and H. bulbosum (2x=14; 4x=28) were estimated on the basis of the karyotypes and the pairing behaviour of the chromosomes in diploid, triploid and tetraploid hybrids obtained with the aid of embryo culture. — A comparison of the karyotypes of the two species revealed similarities as well as differences. It was concluded that at least 4 or more of the chromosomes were similar in morphology and probably closely related. — Diploid and tetraploid hybrids are rarely obtained and their chromosome numbers tend to be unstable whereas triploid hybrids (1 vulgare + 2 bulbosum genomes) were stable and relatively easy to produce. In the diploid hybrid only 40% of the meiotic cells contained 14 chromosomes while the numbers ranged from 7 to 16 in other cells. All hybrids exhibited pairing between the chromosomes of the two species. Diploid hybrids had a mean of 5.0 and a maximum of 7 bivalents per cell in those cells having 14 chromosomes. Triploid hybrids from crosses between 2x H. vulgare and 4x H. bulbosum exhibited a mean of 1.5 and a maximum of 5 trivalents per cell. In a hexaploid sector found following colchicine treatment of a triploid the mean frequencies of chromosome associations per cell were: 5.5_I+8.0_II+0.7_III+3.7_IV+0.3_V+0.4_VI. One unstable 27 chromosome hybrid obtained from crosses between the autotetraploid forms had a mean of 1.1 and a maximum of 4 quadrivalents per cell. The chromosome associations observed in these hybrids are consistent and are taken as evidence of homoeologous pairing between the chromosomes of the two species. Interspecific hybridization between these two species also reveals that chromosome stable hybrids are only obtained when the genomes are present in a ratio of 1 vulgare2 bulbosum. Based upon the results obtained, the possibility of transferring genetic characters from H. bulbosum into cultivated barley is discussed.     

Numerical analysis of isozyme variation in Glycine tomentella

Isozyme variation among 114 accessions of the Glycine tomentella Hayata was analysed by single linkage cluster analysis and the unweighted pair group centroid method (UPGMC). The diploid accessions fell into six distinct, well defined groups, which conformed with differences in chromosome number (2n − 38 or 2n − 40) or in geographic origin. The majority of the tetraploid accessions belonged to a large, geographically widespread group, predominantly aneuploid (2n − 78) group. The remaining four tetraploid groups were distinct on the basis of morphology or geographic distribution. The validity of tetraploid isozyme groupings for reflecting subspecific differentiation was supported by the published reports of hybrid fertility. All of the nineteen crosses between isozyme groups have yielded sterile hybrids, whereas five crosses within groups have yielded fertile hybrids. The relationship between diploids and tetraploids was examined either as the similarity between individual accessions, or that between isozyme groups. These analyses indicated that each tetraploid group is closely related to only one or two of the diploid groups or subgroups.     

Plant regeneration from seedling explants of perennial Glycine species

More than 5000 cultures, from 30 accessions of six Glycine species, were established to assess the rôle of plant genotype in the response to an agar-solidified culture medium containing B5 salts and vitamins, 3% w/v sucrose, 1.1 mg 1^–1 BAP and 0.005 mg 1^–1 IBA, already known to induce shoot regeneration in callus of G. clandestina. Shoot initiation was obtained in a variety of explants from G. canescens, G. falcata, G. latrobeana and G. tomentella. With the exception of G. latrobeana, development of buds into shoots followed transfer to B5-based medium with 0.2 mg^–1 BAP and 0.005 mg 1^–1 IBA. Shoots readily produced roots in hormone-free half-strength B5 medium. In G. latrobeana, both extension and rooting occurred on this medium. Shoot regeneration was obtained in 12 of 30 accessions evaluated, but one accession of G. canescens, G1171, produced shoots and plantlets at a consistently higher frequency than other accessions, with plantlet recovery in more than 70% of the cultures. Bud formation in callus of G. canescens G1171 also occurred if BAP was replaced by 1.0 mg 1^–1 kinetin, 2i-p or zeatin, albeit at a lower frequency.