Quantitative trait loci and candidate gene mapping of aluminum tolerance in diploid alfalfa.

Aluminum (Al) toxicity in acid soils is a major limitation to the production of alfalfa (Medicago sativa subsp. sativa L.) in the USA. Developing Al-tolerant alfalfa cultivars is one approach to overcome this constraint. Accessions of wild diploid alfalfa (M. sativa subsp. coerulea) have been found to be a source of useful genes for Al tolerance. Previously, two genomic regions associated with Al tolerance were identified in this diploid species using restriction fragment length polymorphism (RFLP) markers and single marker analysis. This study was conducted to identify additional Al-tolerance quantitative trait loci (QTLs); to identify simple sequence repeat (SSR) markers that flank the previously identified QTLs; to map candidate genes associated with Al tolerance from other plant species; and to test for co-localization with mapped QTLs. A genetic linkage map was constructed using EST-SSR markers in a population of 130 BC(1)F(1) plants derived from the cross between Al-sensitive and Al-tolerant genotypes. Three putative QTLs on linkage groups LG I, LG II and LG III, explaining 38, 16 and 27% of the phenotypic variation, respectively, were identified. Six candidate gene markers designed from Medicago truncatula ESTs that showed homology to known Al-tolerance genes identified in other plant species were placed on the QTL map. A marker designed from a candidate gene involved in malic acid release mapped near a marginally significant QTL (LOD 2.83) on LG I. The SSR markers flanking these QTLs will be useful for transferring them to cultivated alfalfa via marker-assisted selection and for pyramiding Al tolerance QTLs.

Investigation of genomic organization in switchgrass (Panicum virgatum L.) using DNA markers.

We report an early investigation into genomic organization and chromosomal transmission in switchgrass based on restriction fragment length polymorphism (RFLP) markers. The segregation of 224 single dose restriction fragments (SDRF) in 85 full-sib progeny of a cross between the genotypes Alamo (AP13) and Summer (VS16) was used to determine linkage associations in each parent. In the seed parent AP13, 11 cosegregation groups were identified by 45 SDRF markers with a cumulative recombination length of 412.4 cM. In the pollen parent VS16, 57 SDRF markers were assigned to 16 cosegregation groups covering a length of 466.5 cM. SDRF markers identified by the same probes and mapping to different cosegregation groups were used to combine the two maps and identify homology groups. Eight homology groups were identified among the nine haploid linkage groups expected in switchgrass. The high incidence of repulsion phase associations indicates that preferential pairing between homologous chromosomes is predominant in switchgrass. Based on marker distribution in the paternal map (VS16), we estimated the recombinational length of switchgrass genome to be 4,617 cM. In order to link 95% of the genome to a marker at a 15-cM distance, a minimum of 459 markers will be required. Using information from the ratio of repulsion to coupling linkages, we infer that switchgrass is an autotetraploid with a high degree of preferential pairing. The information presented in this study establishes a foundation for extending genetic mapping in this crop and constitutes a framework for basic and applied genetic studies.

Productivity of cow-calf pairs grazing tall fescue pastures infected with either the wild-type endophyte or a nonergot alkaloid-producing endophyte strain, AR542.

The nonergot alkaloid-producing endo-phyte, AR542, has been shown to improve the persistence and yield of tall fescue pastures without causing the animal disorders commonly associated with tall fescue toxicosis. A 3-yr grazing study was conducted to compare effects of AR542-infected tall fescue pastures with wild type endophyte-infected (E+) tall fescue pastures on cow-calf performance. Replicated 7.3-ha pastures of each treatment were grazed by cow-calf pairs (16 pairs per pasture replication) each year from March to weaning in September. The cows were exposed to breeding on their respective pasture treatments from April 1 through June 15. The treatment groups were compared for reproductive performance, ADG, BCS, calf growth rate, and weaning weight. Blood samples were also collected for serum prolactin (PRL) analysis. There were no significant differences in calving rate (P = 0.98) or calving interval (P = 0.62) between pasture treatments. Cows that grazed the AR542 pastures subsequently gave birth to calves that were heavier (P < 0.05) than calves from cows that had grazed the E+ pastures. Cows grazing the AR542 pastures had higher (P < 0.05) BCS at the end of the grazing period, and had higher ADG during the grazing period. Calves raised on the AR542 pasture had higher (P < 0.05) ADG and weaning weights than calves of the same sex raised on the E+ pastures. Serum PRL concentrations were decreased (P < 0.05) in both cows and calves on the E+ pastures compared with serum PRL concentrations in cows and calves grazing the AR542 pastures. The results indicate that grazing tall fescue pastures infected with the AR542 endophyte may give significant advantages in cow-calf growth rates and BCS over grazing E+ pastures. However, there did not seem to be any benefit in reproductive performance in this trial. There was a small, but significant increase in birth weight in cows grazing AR542 pasture.

Use of nonergot alkaloid-producing endophytes for alleviating tall fescue toxicosis in stocker cattle.

Grazing studies were conducted to determine cattle growth performance, evaluate toxicosis, and compare grazing behavior in stocker cattle grazing nonergot alkaloid-producing endophyte-infected (AR542 or AR502), endophyte-free (E-), or wild-type toxic endophyte-infected (E+) Jesup, Georgia-5, and Kentucky-31 tall fescue. Replicated 0.81-ha tall fescue paddocks were established at the Central Georgia Branch Station at Eatonton and the Northwest Georgia Branch Station at Calhoun during October 1998 and were stocked with beef cattle for autumn and spring periods from fall 1999 through spring 2002. Mean ergot alkaloid concentrations were higher (P < 0.01) on E+ pastures than the other treatments at both locations. At Calhoun and Eatonton, post-treatment serum prolactin concentrations were decreased (P < 0.01) on E+ compared with AR542, AR502, and E- tall fescue. Cattle on AR542, AR502, and E- pastures had lower (P < 0.05) post-treatment rectal temperatures than cattle grazing E+ tall fescue during spring at Eatonton and Calhoun. Calf ADG was higher (P < 0.05) on AR542, AR502, and E- as compared with E+ tall fescue during autumn and spring grazing at Eatonton, and at Calhoun, cattle on E+ pastures had lower (P < 0.05) ADG in both autumn and spring. Gain/hectare was higher (P < 0.05) on AR542, AR502, and E- than on E+ during autumn at Eatonton and during spring at both locations. In autumn at Calhoun, gain/hectare was greater (P < 0.05) on AR502 and E- compared with E+ tall fescue. During April, May, and June, cattle grazing E+ pastures at Eatonton spent more (P < 0.01) time idling, more (P < 0.01) time standing, and used more (P < 0.01) water than cattle on AR542 and E- tall fescue. Daily prehensions and biting rate were each higher (P < 0.01) on AR542 and E- tall fescue than E+ tall fescue in both grazing seasons. There were no differences among pasture treatments for bite size in either spring (P = 0.50) or autumn (P = 0.34). Steers grazing E+ pastures had lower DMI than steers grazing AR542 and E- pastures during spring (P < 0.10) and lower DMI than steers grazing E- pastures during autumn (P < 0.05). Daily steer water usage was decreased (P < 0.10) in E+ pastures compared with AR542 and E- pastures during late fall. These results indicate that nonergot alkaloid-producing endophyte technology is a promising option for alleviating tall fescue toxicosis in stocker cattle.

Use of nonergot alkaloid-producing endophytes for alleviating tall fescue toxicosis in sheep.

Nonergot alkaloid-producing endophytes from New Zealand were inserted into tall fescue (Festuca arundinacea) cultivars in an attempt to address the problem of fescue toxicosis in grazing sheep. A 3-yr grazing study was conducted to determine lamb performance and to evaluate toxicosis in lambs grazing nonergot alkaloid-producing endophyte-infected (AR542 or AR502), endophyte-free (E-), or wild-type toxic endophyte-infected (E+) Jesup tall fescue or nonergot alkaloid-producing endophyte-infected (AR542) Georgia-5 tall fescue. Replicated 0.11-ha tall fescue paddocks were established at the central Georgia Branch Station during September 1997 and stocked with lambs from spring 1998 through autumn 2000. Mean ergot alkaloid concentrations were higher (P < 0.01) in E+ forage than in AR542, AR502, and E- tall fescue, and ergot alkaloid concentrations in E- plants and plants infected with AR542 and AR502 were low. Forage availability did not differ (P = 0.92) across treatments during autumn and was higher (P < 0.05) in Georgia-5 AR542 than in Jesup AR502 and E+ pastures. Initial serum prolactin (PRL) concentrations did not differ (P = 0.58) across treatments during autumn, but were higher on Jesup AR542 than E+ during spring. Post-treatment serum PRL concentrations were depressed (P < 0.01) on E+ compared with AR542, AR502, and E- in both spring and autumn. Signs of heat stress were observed in E+ lambs during periods of high ambient temperatures. Mean post-treatment rectal temperature and mean stocking rate exhibited treatment x year interactions (P < 0.05). Lamb ADG was higher (P < 0.05) on AR542, AR502, and E- than on E+ tall fescue. Similarly, gain/hectare was higher (P < 0.015) on AR542, AR502, and E- than on E+. Tall fescue pastures containing AR542 and AR502 endophytes yielded lamb performance that did not differ from that on E- tall fescue and which was superior to performance on E+ tall fescue. Depressed PRL concentrations and elevated rectal temperatures as indicators of toxicosis were evident only in lambs grazing E+ tall fescue, suggesting that nonergot alkaloid-producing endophyte-infected tall fescue is a viable alternative for alleviating tall fescue toxicosis.

Alfalfa weevil (Coleoptera:Curculionidae) management in alfalfa by spring grazing with cattle.

The effect of continuous, intensive grazing by cattle in the 1st alfalfa growth cycle on larval densities of the alfalfa weevil, Hyera postica (Gyllenhal), was evaluated in "Alfagraze' and "Apollo' alfalfa, which are tolerant and not tolerant to grazing, respectively. In small-cage exclusion trials, grazing reduced larval numbers in 1991 by 65% in Alfagraze and by 32% in Apollo. Larval numbers in 1992 were low (< or = 0.6 larvae per stem) and were not reduced significantly by grazing. Grazing and use of early insecticide treatments of permethrin or carbofuran at low rates with < or = 7-d grazing restrictions to suppress larval numbers before grazing also were examined in large-plot exclusion trails in 1993 and 1994. Grazing reduced larval densities by 60% in 1993 and 45% in 1994 during a 3-wk period beginning 3 wk after grazing was initiated. However, alfalfa weevil larvae caused moderate leaf injury in 1993 and severe injury in 1994 before grazing reduced larval numbers. Use of permethrin at 0.11 kg (AI)/ha or carbofuran or chlorpyrifos at 0.28 kg (AI)/ha effectively reduced larval numbers and prevented leaf injury before grazing began. Therefore, a combination of an early application of an insecticide treatment with a short grazing restriction followed by continuous grazing will control alfalfa weevil larvae while allowing cattle to graze and directly use forage of grazing-tolerant alfalfa.

Analysis of annual Medicago species using RAPD markers.

Annual species of the genus Medicago have attracted interest as green manure and temporary forage crops. This study was conducted to determine if randomly amplified polymorphic DNA (RAPD) markers could be used to assess the variability within and among species. Several accessions of each six species (M. scutellata Mill., M. disciformis DC., M. murex Willd., M. truncatula Gaertn., M. polymorpha L., and M. rugosa Desr.) were studied. A phylogeny reconstructed with the computer program Phylogenetic Analysis Using Parsimony (PAUP) showed the same relationships as traditional taxonomy. Variation was present among accessions of all species. Several accessions were considerably different from others within the species (one of each M. scutellata and M. polymorpha) and four accessions of M. murex were differentiated by both morphology and RAPD banding patterns from the other accessions. These accessions may be useful to include in a core collection. Variation within accessions was present. Although the species are autogamous, the original seed collections may have been made from a number of plants in the same area. Also, some outcrossing or seed mixing may have occurred. Finally, at least 10 RAPD primers appear to be necessary in order to develop reliable estimates of relatedness among annual Medicago accessions.

Photosynthetic Characteristics of Segregates from Hybrids between Flaveria brownii (C4 Like) and Flaveria linearis (C3-C4).

Characteristics related to C4 photosynthesis were studied in reciprocal F1 hybrids and F2 plants from Flaveria brownii (C4 like) and Flaveria linearis (C3-C4). The reciprocal F1 plants differed in 13C/12C ratios of leaves and the percentage of 14C initially incorporated into C4 acids, being more like the pollen parents in these traits. They did not differ in apparent photosynthesis or in O2 inhibition of apparent photosynthesis and differed only slightly in CO2 compensation concentration at 175 [mu]mol quanta m-2 s-1 and 400 mL L-1 O2. The 13C/12C ratios of 78 F2 progeny from the two F1 plants exhibited a normal distribution centered between those of the parents, with a few values slightly higher and lower than the parents. Apparent photosynthesis at 130 [mu]L L-1 CO2 and inhibition of photosynthesis by O2 was nearly normally distributed in the F2 population, but no values for F2 plants approached those for F. brownii (15.4 [mu]mol m-2 s-1 and 7.8%, respectively). Distribution of the CO2 compensation concentration measured at 1000 [mu]mol quanta m-2 s-1 and 400 mL L-1 of O2 in the F2 population was skewed toward F. brownii with 72% of the progeny having values <9 [mu]L of CO2 L-1 compared to 1.5 and 27.2 [mu]L L-1 for F. brownii and F. linearis, respectively. Correlations among traits of F2 plants were low (coefficients of 0.30 to -0.49), indicating that the C4- related traits are not closely linked in segregating populations. Plants in the F2 population selected for high or low apparent photosynthesis at 130 [mu]L of CO2 L-1 (six each) did not rank consistently high or low for 13C/12C ratios, O2 inhibition of apparent photosynthesis, CO2 compensation concentration, or activities of phosphoenolpyruvate carboxylase or NADP-malic enzyme. This study confirms results of earlier work that indicates independent segregation of C4 traits and also shows that the C4-like parental type can be recovered, at least for some characteristics (13C/12C ratio), in segregating populations. Recovery of fully functional C4 plants awaits further experimentation with C4 x C3 or C4 x C3-C4 hybrid plants that produce fertile progeny.

Development of an RFLP map in diploid alfalfa.

We have developed a restriction fragment length polymorphism (RFLP) linkage map in diploid alfalfa (Medicago sativa L.) to be used as a tool in alfalfa improvement programs. An F2 mapping population of 86 individuals was produced from a cross between a plant of the W2xiso population (M. sativa ssp. sativa) and a plant from USDA PI440501 (M. sativa ssp. coerulea). The current map contains 108 cDNA markers covering 467.5 centimorgans. The short length of the map is probably due to low recombination in this cross. Marker order may be maintained in other populations even though the distance between clones may change. About 50% of the mapped loci showed segregation distortion, mostly toward excess heterozygotes. This is circumstantial evidence supporting the maximum heterozygote theory which states that relative vigor is dependent on maximizing the number of loci with multiple alleles. The application of the map to tetraploid populations is discussed.

Degree of C(4) Photosynthesis in C(4) and C(3)-C(4)Flaveria Species and Their Hybrids : I. CO(2) Assimilation and Metabolism and Activities of Phosphoenolpyruvate Carboxylase and NADP-Malic Enzyme.

The degree of C(4) photosynthesis was assessed in four hybrids among C(4), C(4)-like, and C(3)-C(4) species in the genus Flaveria using (14)C labeling, CO(2) exchange, (13)C discrimination, and C(4) enzyme activities. The hybrids incorporated from 57 to 88% of the (14)C assimilated in a 10-s exposure into C(4) acids compared with 26% for the C(3)-C(4) species Flaveria linearis, 91% for the C(4) species Flaveria trinervia, and 87% for the C(4)-like Flaveria brownii. Those plants with high percentages of (14)C initially fixed into C(4) acids also metabolized the C(4) acids quickly, and the percentage of (14)C in 3-phosphoglyceric acid plus sugar phosphates increased for at least a 30-s exposure to (12)CO(2). This indicated a high degree of coordination between the carbon accumulation and reduction phases of the C(4) and C(3) cycles. Synthesis and metabolism of C(4) acids by the species and their hybrids were highly and linearly correlated with discrimination against (13)C. The relationship of (13)C discrimination or (14)C metabolism to O(2) inhibition of photosynthesis was curvilinear, changing more rapidly at C(4)-like values of (14)C metabolism and (13)C discrimination. Incorporation of initial (14)C into C(4) acids showed a biphasic increase with increased activities of phosphoenolpyruvate carboxylase and NADP-malic enzyme (steep at low activities), but turnover of C(4) acids was linearly related to NADP-malic enzyme activity. Several other traits were closely related to the in vitro activity of NADP-malic enzyme but not phosphoenolpyruvate carboxylase. The data indicate that the hybrids have variable degrees of C(4) photosynthesis but that the carbon accumulation and reduction portions of the C(4) and C(3) cycles are well coordinated.

RFLP variation in diploid and tetraploid alfalfa.

Alfalfa (Medicago sativa L.) is a major forage crop throughout the world. Although alfalfa has many desirable traits, continued breeding is required to incorporate pest resistances and other traits. We conducted this study to determine the amount of restriction fragment length polymorphism (RFLP) variability present within and between diploid and tetraploid alfalfa populations, and whether or not this variability is sufficient for construction of an RFLP map. Diploid plants from M. sativa ssp. falcata, ssp. coerulea, and ssp. sativa and tetraploid spp. sativa cultivars 'Apollo,' 'Florida 77,' and 'Spredor 2' were included. A total of 19 cDNA clones was probed onto genomic Southern blots containing DNA digested by EcoRI, HindIII, or BamHI. Phylogenetic trees were produced, based on parsimony analysis of shared restriction fragments. Evidence for extensive gene duplication was found; most probes detected complex patterns of restriction fragments. Large amounts of variation are present within all diploid subspecies. M. sativa ssp. falcata plants formed clusters distinct from ssp. sativa or ssp. coerulea plants, which were not distinctly clustered. Some M. sativa ssp. falcata plants were more similar to the other groups than to other plants within ssp. falcata. Variation among tetraploid cultivars showed that Florida 77 and Apollo had more similarities than either showed with Spredor 2. All three cultivars showed large within-population variation, with Apollo being the most diverse and Spredor 2 the least. Based on these results, development of an RFLP map at the diploid level appears possible. Also, differentiation of cultivars, particularly ones of divergent origin, seems possible based on RFLP patterns.

Inheritance of the Reversal of O(2) Response of Photosynthesis in a Flaveria linearis Mutant.

A mutant plant of Flaveria linearis Lag. expresses reversed O(2) response of photosynthesis (i.e. its apparent photosynthesis is stimulated at atmospheric O(2) levels). The objectives of this study were to determine the genetic inheritance of this trait and to investigate the biochemical mechanism for its expression. The mutant plant was crossed reciprocally with a plant of the closely related species Flaveria oppositifolia (DC.) Rydb. and also with another plant of F. linearis. Data on O(2) inhibition of apparent photosynthesis were analyzed on F(2) and F(3) progeny from these F(1) hybrids. In addition, test crosses (mutant x F(1) hybrid) and S(1) progeny from the mutant plant were also analyzed. All F(1) hybrids expressed inhibition of apparent photosynthesis and their progeny segregated in acceptable 3:1 and 13:3 (normal:reversed) ratios. There was little effect of environment on expression of the reversed O(2) response. Selected F(2) plants and the original mutant plant produced progeny in normal:reversed ratios which indicated the trait is controlled by two major genes which show dominant and recessive epistasis. Plants with greater than 20 nanomoles per gram fresh weight per minute of fructose-1, 6-bisphosphatase activity in the cytosol had normal O(2) response of photosynthesis. However, when plants had less than 20 nanomoles per gram fresh weight per minute of this enzyme activity in the cytosol, the O(2) was normal in some and reversed in others. It is proposed that low fructose bisphosphatase activity in the cytosol is controlled by a recessive gene (fbp). A second dominant gene is speculated to be hypostatic to the normal fructose bisphosphatase gene and controls the expression of an unknown factor that determines whether O(2) response of AP is reversed in the presence of fbp (i.e. when fructose bisphosphatase activity is low).

Leaf anatomical characteristics in Flaveria trinervia (C4), Flaveria brownii (C 4-like) and their F 1 hybrid.

Several leaf anatomical and ultrastructural characteristics usually related with photosynthetic capacity were examined in two Flaveria species with strong differences in anatomy and their F1 hybrid. Flaveria trinervia (Spreng.) Mohr (C4) was the female parent and F. brownii A.M. Powell (C4-like) was the male parent. Quantitative anatomical analysis was made on transverse sections of leaves at both the light and electron microscope level. Four kinds of photosynthetic tissues were considered: bundle sheath (BS), mesophyll adjacent to the BS, mesophyll not adjacent to the BS, and larger spongy mesophyll cells. Flaveria trinvervia partitioned a larger proportion of its photosynthetic cells to BS and the mesophyll layer adjacent to BS and also possessed larger chloroplasts, especially in BS, than did F. brownii. These results suggest that although F. brownii is very C4-like, its anatomy is not as completely C4 as is the case for F. trinervia. In the F1 hybrid the relative contribution of the different tissues to the total photosynthetic tissue volume and area per unit leaf area was quite similar to that of F. trinervia. On the other hand, the chloroplast density and size of the F1 hybrid were fairly similar to those of F. brownii, especially in BS. Thus, there was no evidence of maternal inheritance in the chloroplast characteristics studied. A negative correlation (P<0.05) between chloroplast size and density was observed among species and relicates within each kind of tissue. This correlation was highest (r=-0.94, P<0.001) for the BS and when values were plotted on a logarithmic scale. Thus, higher chloroplast numbers for F. brownii and the F1 hybrid were offset by larger chloroplasts in F. trinervia. Less complete C4 photosynthesis in F. brownii may be partially due to incomplete development of Kranz anatomy usually associated with C4 photosynthesis.

Transfer of C(4) Photosynthetic Characters through Hybridization of Flaveria Species.

Transfer of C(4) photosynthetic traits was studied through hybridization of Flaveria trinervia (Spreng.) Mohr (C(4)) and Flaveria brownii A.M. Powell (C(4)-like) with Flaveria linearis Lag. (C(3)-C(4)) and the C(3) species Flaveria pringlei Gandoger (C(3)). Fertility was low, based on irregular chromosome pairing and low pollen stainability, except in F. brownii x F. linearis which had bivalent pairing and 76% stainable pollen. Hybrids had apparent photosynthesis values of 71 to 148% of the midparental means, while the CO(2) compensation concentration was similar to the C(4) or C(4)-like parent, except in hybrids having the C(3) species F. pringlei as a parent. Inhibition of apparent photosynthesis by O(2), and phosphoenolpyruvate carboxylase and NADP-malic enzyme activities and subunit levels in the hybrids were closer to the C(3) or C(3)-C(4) parent. The species F. brownii and F. trinervia were equal in their capacity to transfer reduced O(2) inhibition of AP and CO(2) compensation concentration values to hybrids with F. linearis (C(3)-C(4)), although hybrids with F. trinervia had higher PEPC activity. The O(2) inhibition of AP was correlated with the logarithm of activities of phosphoenolpyruvate carboxylase (r = -0.95) and NADP-malic enzyme (r = -0.87). These results confirm that C(4) traits can be transferred by hybridization of C(3)-C(4) and C(4) or C(4)-like species, with a higher degree of C(4) photosynthesis than exists in C(3)-C(4) species, and at least in F. brownii x F. linearis, fertile progeny are obtained.

CO(2) Exchange, Cytogenetics, and Leaf Anatomy of Hybrids between Photosynthetically Distinct Flaveria Species.

Hybrids between the C(4)-like species, Flaveria brownii, A. M. Powell and the C(3)-C(4) intermediate species Flaveria linearis Lag., Flaveria floridana Johnston, and Flaveria oppositifolia (DC.) Rydb. exhibited bivalent chromosome pairing during meiosis and stainability of pollen was high, ranging from 51 to 95%. An F(2) population produced from an F. brownii x F. linearis F(1) hybrid, exhibited bivalent chromosome pairing and high pollen stainability indicating a high degree of fertility in the hybrid. Oxygen inhibition of apparent photosynthesis averaged 6.8% for F. brownii and 22.2% for the C(3)-C(4) species (in two experiments), and F(1) hybrids exhibited inhibitions which were intermediate to their parents. Values of carbon dioxide compensation concentration determined at low irradiance were 4.0, 34.0, and 6.5 microliters per liter for F. brownii, F. linearis and their F(1) hybrid, respectively. The mean value at low irradiance for 33 F(1) plants was 6.8 microliters per liter, and individual values ranged only from 3.7 to 11.7 microliters per liter. Anatomical characteristics for the F(1) hybrid leaves were intermediate to those of the parents, and there was considerable variation among F(2) plants derived from F. brownii x F. linearis. In the F(2) population delta(13)C values ranged from -27 per thousand to -20 per thousand. The expression of more C(4)-like characteristics by the F(1) hybrids in this study and their apparent high fertility make them promising specimens for producing segregating populations for use in C(4) inheritance studies.

Photosynthesis of F(1) Hybrids between C(4) and C(3)-C(4) Species of Flaveria.

Photosynthetic characteristics were studied in several F(1) hybrids between C(4) and C(3)-C(4) species of Flaveria. Stable carbon isotope ratios, O(2) inhibition of apparent photosynthesis, and phosphoenolpyruvate carboxylase activities in the hybrids were similar to the means for the parents. Values of CO(2) compensation concentrations were nearer to those of the C(4) parent and apparent photosynthesis was below that of both parents, being only 60 and 74% of that of the lowest (C(3)-C(4)) parent in two experiments. Reductions of CO(2) compensation concentration and O(2) inhibition of apparent photosynthesis as well as increases in carbon isotope ratios and phosphoenolpyruvate carboxylase activities compared to values in C(3)-C(4) species suggest transfer of a limited degree of C(4) photosynthesis to the F(1) hybrids. However, the lower apparent photosynthesis of the hybrids suggests that transfer of C(4) characteristics to non-C(4) species is detrimental unless characteristics associated with C(4) photosynthesis are fully developed. There was a highly significant negative correlation (r = -0.90) between CO(2) compensation concentration and the logarithm of phosphoenolpyruvate carboxylase activity in the parents and hybrids, suggesting involvement of this enzyme in controlling the CO(2) compensation concentration. Although bundle-sheath cells were more developed in leaves of hybrids than in C(3)-C(4) parents, they appeared to contain lower quantities of organelles than those of the C(4) parent. Reduced quantities of organelles in bundle-sheath cells could indicate incomplete compartmentation of partial pathways of the C(4) cycle in the hybrids. This may mean that the reduction of CO(2) compensation and O(2) inhibition of apparent photosynthesis relative to the C(3)-C(4) parents is less dependent on fully developed Kranz anatomy than is increased apparent photosynthesis.

Oxygen Stimulation of Apparent Photosynthesis in Flaveria linearis.

A plant was found in the C(3)-C(4) intermediate species, Flaveria linearis, in which apparent photosynthesis is stimulated by atmospheric O(2) concentrations. A survey of 44 selfed progeny of the plant showed that the O(2) stimulation of apparent photosynthesis was passed on to the progeny. When leaves equilibrated at 210 milliliters per liter O(2) were transferred to 20 milliliters per liter O(2) apparent photosynthesis was initially stimulated, but gradually declined so that at 30 to 40 minutes the rate was only about 80 to 85% of that at 210 milliliters per liter O(2). Switching from 20 to 210 milliliters per liter caused the opposite transition in apparent photosynthesis. All other plants of F. linearis reached steady rates within 5 minutes after switching O(2) that were 20 to 24% lower in 210 than in 20 milliliters per liter O(2). At low intercellular CO(2) concentrations and low irradiances, O(2) inhibition of apparent photosynthesis of the aberrant plant was similar to that in normal plants, but at an irradiance of 2 millimoles quanta per square meter per second and near 300 microliters per liter CO(2) apparent photosynthesis was consistently higher at 210 than at 20 milliliters per liter O(2). In morphology and leaf anatomy, the aberrant plant is like the normal plants in F. linearis. The stimulation of apparent photosynthesis at air levels of O(2) in the aberrant plant is similar to other literature reports on observations with C(3) plants at high CO(2) concentrations, high irradiance and/or low temperatures, and may be related to limitation of photosynthesis by triose phosphate utilization.

Photosynthesis, Leaf Anatomy, and Morphology of Progeny from Hybrids between C(3) and C(3)/C(4)Panicum Species.

Species in the Laxa group of Panicum have C(3) or C(3)/C(4) photosynthesis based on leaf anatomical and CO(2) exchange characteristics. Hybrids were previously made between C(3)/C(4) and C(3) species in this group (RH Brown et al. 1985 Plant Physiol 77: 653-658). In this paper, CO(2) exchange, morphological, and leaf anatomical characteristics of F(2) or F(5) progeny from colchicine-induced amphiploids of C(3)/C(4) x C(3) hybrids (Panicum milioides Nees ex Trin. [C(3)/C(4)] x Panicum laxum Mez [C(3)] and Panicum spathellosum Doell [C(3)/C(4)] x Panicum boliviense Hack. [C(3)]) were studied.There were no differences found in morphology or physiology between the amphiploids and the F(1) hybrids from which they were produced. In the segregating progeny, CO(2) compensation concentration and photorespiration values typical of C(3), but not of C(3)/C(4) plants, were recovered. Progeny were found from both crosses which possessed O(2) inhibition of apparent photosynthesis typical of the parents, and in the case of the P. milioides x P. laxum cross, leaf anatomy and overall plant morphology typical of the parents were observed in some progeny. The progeny were found to possess recombinations of various traits associated with reduced photorespiration, so that no correlation existed among O(2) inhibition of apparent photosynthesis, CO(2) compensation concentration, and leaf anatomical traits. One plant was especially noteworthy in possessing leaf anatomy typical of C(3)/C(4) plants, but with CO(2) exchange characteristics of C(3) plants.

Photosynthesis, Morphology, Leaf Anatomy, and Cytogenetics of Hybrids between C(3) and C(3)/C(4)Panicum Species.

The Laxa group of the Panicum genus contains species which have CO(2) exchange and anatomical characteristics intermediate to C(3) and C(4) photosynthetic types (C(3)/C(4)), and also species characterized as C(3). Hybrids were made between two of the C(3)/C(4) species and two C(3) species. Carbon dioxide exchange and morphological, leaf anatomical, and cytogenetic characteristics of F(1) hybrids between Panicum milioides Nees. ex Trin (C(3)/C(4)) and P. laxum Mez. (C(3)), P. spathellosum Doell (C(3)/C(4)) and P. boliviense Hack. (C(3)), and P. spathellosum and P. laxum were studied. There were no consistent differences in apparent photosynthesis, although two of the three hybrids had higher net CO(2) uptake than the C(3) parent. Values of inhibition of apparent photosynthesis by 21% O(2), CO(2) loss in the light, and CO(2) compensation concentration for the hybrids were between those of the parents. All three hybrids showed leaf anatomical traits, especially organelle quantities in the bundle sheath cells, between those of their respective parents. Linear regression of CO(2) compensation concentration on the percentage of mitochondria and chloroplasts in vascular bundle sheaths of the parents and hybrids gave correlation coefficients of -0.94. This suggests that the reduction in CO(2) loss in the C(3)/C(4) species, and to a lesser degree in the F(1) hybrids, was due to development of organelles and perhaps a higher proportion of leaf photorespiration in bundle sheaths. The overall morphology of the hybrids was so different from the parents that they could be described as new taxonomic forms. The chromosomes in the hybrids were mainly unpaired or paired as bivalents indicating possible homology between some parental genomes.

Photosynthesis of Grass Species Differing in Carbon Dioxide Fixation Pathways : VIII. Ultrastructural Characteristics of Panicum Species in the Laxa Group.

Ultrastructural studies of leaves of seven Panicum species in or closely related to the Laxa group and classified as C(3), C(4) or C(3)-C(4) intermediate were undertaken to examine features associated with C(3) and C(4) photosynthesis. The C(3) species Panicum rivulare Trin. had few organelles in bundle sheath cell profiles (2 chloroplasts, 1.1 mitochondria, and 0.3 peroxisomes per cell section) compared to an average of 10.6 chloroplasts, 17.7 mitochondria, and 3.2 peroxisomes per bundle sheath cell profile for three C(3)-C(4) species, Panicum milioides Nees ex Trin., Panicum decipiens Nees ex Trin. and Panicum schenckii Hack. However, two other C(3) species, Panicum laxum Sw. and Panicum hylaeicum Mez, contained about 0.7, 0.5, and 0.3 as many chloroplasts, mitochondria, and peroxisomes, respectively, as in bundle sheath cell profiles of the C(3)-C(4) species. Chloroplasts and mitochondria in bundle sheath cells were larger than those in mesophyll cells for the C(4) species Panicum prionitis Griseb. and the C(3)-C(4) species, but in C(3) species the organelles were similar in size or were smaller in the bundle sheath cells. The C(3)-C(4) species and P. laxum and P. hylaeicum exhibited an unusually close association of organelles in bundle sheath cells with mitochondria frequently surrounded in profile by chloroplasts. The high concentrations in bundle sheath cells of somewhat larger organelles than in mesophyll cells correlates with the reduced photorespiration of the C(3)-C(4) species.