Molecular diversity and association mapping of fiber quality traits in exotic G. hirsutum L. germplasm.

The narrow genetic base of cultivated cotton germplasm is hindering the cotton productivity worldwide. Although potential genetic diversity exists in Gossypium genus, it is largely 'underutilized' due to photoperiodism and the lack of innovative tools to overcome such challenges. The application of linkage disequilibrium (LD)-based association mapping is an alternative powerful molecular tool to dissect and exploit the natural genetic diversity conserved within cotton germplasm collections, greatly accelerating still 'lagging' cotton marker-assisted selection (MAS) programs. However, the extent of genome-wide linkage disequilibrium (LD) has not been determined in cotton. We report the extent of genome-wide LD and association mapping of fiber quality traits by using a 95 core set of microsatellite markers in a total of 285 exotic Gossypium hirsutum accessions, comprising of 208 landrace stocks and 77 photoperiodic variety accessions. We demonstrated the existence of useful genetic diversity within exotic cotton germplasm. In this germplasm set, 11-12% of SSR loci pairs revealed a significant LD. At the significance threshold (r(2)>/=0.1), a genome-wide average of LD declines within the genetic distance at <10 cM in the landrace stocks germplasm and >30 cM in variety germplasm. Genome wide LD at r(2)>/=0.2 was reduced on average to approximately 1-2 cM in the landrace stock germplasm and 6-8 cM in variety germplasm, providing evidence of the potential for association mapping of agronomically important traits in cotton. We observed significant population structure and relatedness in assayed germplasm. Consequently, the application of the mixed liner model (MLM), considering both kinship (K) and population structure (Q) detected between 6% and 13% of SSR markers associated with the main fiber quality traits in cotton. Our results highlight for the first time the feasibility and potential of association mapping, with consideration of the population structure and stratification existing in cotton germplasm resources. The number of SSR markers associated with fiber quality traits in diverse cotton germplasm, which broadly covered many historical meiotic events, should be useful to effectively exploit potentially new genetic variation by using MAS programs.

Diallel analysis of superior fiber quality properties in selected upland cottons.

Twenty cross combinations were produced using a complete diallel-mating system with five varieties or lines that differed in fiber properties in Upland cotton to determine the inheritance and breeding merits of superior fiber properties. Evaluations of parents and F1 ' s were conducted in two years. The results showed that fiber length uniformity was greatly affected by environmental factors, whereas the other fiber properties were mainly controlled by genetic factors. There were no significant interaction effect of environment with genotype for fiber strength or length, but there were significant environment interactions with additive and maternal affects for Micronaire, and with the dominance effects for elongation. There were no maternal effect, and additive effects predominated for the all fiber properties. Additive heritability was high for fiber strength and length, 77.6% and 73.2% respectively; for Micronaire, it was 45.2%, while the dominance effect was 11.5%, which was the highest among fiber properties. Micronaire had significant heterosis over mid-parent based on population mean (3.2%), while the other fiber properties showed no heterosis. Therefore, the performance of fiber properties in F1 ' s can be predicted from the average value of both parents. Since the additive heritability of strength, length, and fineness of fiber were high, these traits can be selected in early generations in breeding for high quality fiber properties.

Tests of six cotton (Gossypium hirsutum L.) mutants for association with aneuploids.

Genetic mutants are useful tools for basic and applied research to elucidate the developmental and regulatory processes of the cotton plant (Gossypium hirsutum L.). Their value is enhanced with knowledge of their location in the genome. The results of aneuploid tests used to locate mutant loci on specific chromosomes in G. hirsutum L. are reported. Thirty-four monosomes and telosomes, representing 18 of the 26 chromosomes, were used in combination with six mutants that were associated with nine loci. The mutant loci were glandless stem and boll (gl1gl6), immature fiber (im), Ligon lintless-2 (Li2), methylation (me), nonpinking (np1np2), and Raimondal (Ra1Ra2). We found that im was associated with chromosome 3 that contains linkage group VI (accessory involucre and frego bract); Li2 was associated with chromosome 18 that contains linkage group XVI (open bud and yellow pollen-2); and me was associated with chromosome 9. The remaining three mutants were not associated with the aneuploids in the tests. Knowledge of these chromosome assignments provides a valuable reference for specific studies of mutants and for further genome mapping efforts.

[Molecular tagging and mapping of QTLs for super quality fiber properties in upland cotton].

A G. anomalum introgression line, 7235, characterized as super quality fiber properties, was used to identify molecular markers linked to fiber property QTLs. By use of (7235 x TM-1)F2 in Nanjing and College Station, USA, and (7235 x TM-1)F3 in Nanjing and Hainan. Bulked segregation analysis was employed to produce 3 pairs of mixed DNA pools for fiber strength, micronaire and fiber length according to individual value of (7235 x TM-1)F2 and F2:3. A total of 221 pairs of SSR primers, 1,840 arbitrary 10-mer oligonucleotide primers and 77 ISSR primers were used to screen polymorphism between two parents, and 3 pairs of bulked DNA pools. Fifteen markers amplified by thirteen primers were identified to be linked with fiber quality QTLs through DNA polymorphism surveying between the parents, and then paired bulked DNAs, and screening the individual plant of (7235 x TM1)F2. Linkage test indicated 15 markers could be mapped to three linkage groups. In the first linkage group, eight markers (two SSR and six RAPD markers) associated with fiber strength were tightly linked with 2.2 cM interval genetic distance on average, and located on chromosome 10 in cotton. Two major QTLs for fiber quality characters were identified. One for fiber strength could explain 35% of the phenotypic variation in F2, and 53.8% in F2:3 at Hainan, which has the greatest single QTL effect of fiber strength and could be identified in all four environments, and tightly linked to 6 RAPD markers and 2 SSR markers with genetic distance no more than 16 cM in chromosome 10, in which FSRl933 is the nearest with the distance no more than 0.6 cM. One QTL linked to FMR1(603) for Micronaire could explain 7.8% of the phenotypic variation in F2, and 25.4% in F2:3 at Hainan, and expressed in all four environments. One QTL linked to FLR1(1550) for fiber length could explain 9.5% of the phenotypic variation in F2:3 at Hainan, very little in other environments. So they can be used in marker-assisted selection in increasing fiber quality of commercial cultivars.

Effect of concentration of MgCl2 on random-amplified DNA polymorphism.

The influence of MgCl2 concentration on products in PCR to generate random-amplified polymorphic DNA (RAPD) was evaluated between two highly polymorphic cotton lines. Varying the concentration of MgCl2 (1.5 to 6.0 mM) in the PCR mixtures resulted in both quantitative differences in RAPD PCR-derived DNA bands and qualitative changes in the DNA band patterns. The optimum concentration of MgCl2 was within the range of 1.5 to 4.5 mM in the PCR mixtures for 154 of 200 random primers tested. The optimum concentration of MgCl2 for each primer was independent of the GC/AT ratio of the primer and the number of DNA band(s) amplified.

Maternal effects and generation mean analysis of seed-oil content in cotton (Gossypium hirsutum L.).

The nature of gene action and of maternal influence governing cottonseed oil attributes were determined with four lines, two each with high and low seed-oil percentage. For this purpose, P1, P2, F0, F1, F2 and alternative sets of BC1 and BC2 generations were analysed in six cross-combinations and their reciprocals. Marginal extents of heterosis for seed-oil percentage were noticeable in F1, with inbreeding depression in F2. Data from reciprocal backcrosses provided evidence in favour of maternal rather than cytoplasmic effects of seed-oil development. Relatively higher extents of heterosis, sizeable inbreeding depression and reciprocally unequal F2 averages were characteristic of the seed index trait, which often showed a reversal of effects from F1 to F2. Reverse reciprocal backcrosses exhibited some differences, including greater resemblance between the types, (A/B)A and (B/A)A, in addition to variable dose effects in seed index. Thus, the differences between F1 seed index values were not due to cytoplasmic influence. Positive heterotic effects for seed-oil index, especially among the backcrosses, ranged between 16.08% and 47.29% over midparent averages. Genetic component estimates from analysis of similar sets of crosses differing only in reciprocal backcrosses, and also from sets of reciprocal crosses between any two parental combinations, were inconsistent. Scaling tests detected presence of epistasis within and between a majority of cross-combinations. Despite reciprocal differences, additive gene effects for seed-oil percentage were significant in 7 out of 24 crosses, representing high x low, low x high and low x low seed-oil parents. Those were, however, accompanied by significant dominance effects of higher order. In crosses involving low seed-oil percentage parents SA1060 and SA229, all six components were detected significant, with opposite effects of dominance and dominance x dominance epistatic components. Significant additive components were also detected for seed index and seed-oil index in 7 and 5 out of 24 crosses, respectively. In the inheritance of seed index and seed-oil index, dominance effects were more important. Epistatic components of additive x additive, and to a lesser extent, those of dominant x dominant were found significant.

Enzymic Fractionation of the Stable Carbon Isotopes of Carbon Dioxide by Ribulose-1,5-bisphosphate Carboxylase.

The enzymic fractionation of the stable carbon isotopes of CO(2) (Deltaco(2)) was determined using a purified preparation of ribulose-1,5-bisphosphate (RuBP) carboxylase isolated from cotton (a C(3) plant) leaves. The bicarbonate concentration in the reaction mixture saturated the enzyme and furnished an infinite pool of (12)CO(2) and (13)CO(2) for enzyme fractionation. The RuBP was 96 to 98% pure. The phosphoglycerate synthesized in the reaction mixtures was purified free of RuBP, phosphoglycolate, and other phosphate esters by column chromatography on Dowex 1-Cl(-) resin. The average Deltaco(2) value of -27.1% was determined from five separate experiments. A discussion of the isotope fractionation associated with photosynthetic CO(2) fixation in plants shows that the enzymic fractionation of stable carbon isotopes of CO(2) by RuBP carboxylase is of major importance in determining the delta(13)C values of C(3) plants.

Potential for improving cottonseed quality by genetic and agronomic practices.

Potential utilization of cottonseeds as edible food sources accentuated the need for research on their composition. Studies included evaluation of cottonseed composition; e.g., seed grade, protein, amino acids free fatty acids, oil, fatty acids, cyclopropenoid fatty acids, total gossypol, differential settling as an indicator of potential performance of cottonseed in the liquid cyclone process, and extractability of nonstorage and storage proteins and their gel electrophoretic properties. These extended studies were used to develop a data base on composition of various cottonseed cultivars grown in different locations of Texas that resemble environmentally most of the regions of the United States cotton belt. Tests showed that most constituents of cottonseed vary; statistically significant variables include cultivar, location, and their interaction term, cultivar x location. These data suggest that breeding and agronomic practices could be used to alter cottonseed composition. Although protein quantity of cottonseed from various cultivars differ and can be influenced by agronomic practices, this variability is not reflected in quality of cottonseed protein as detected by gel electrophoretic techniques. Analyses showed that both genetic and agronomic factors influenced formation of edible flour with high protein and low free gossypol content.

Composition and activity of the photosynthetic apparatus in temperature-sensitive mutants of higher plants.

Six nuclear mutants of corn, six of soybean, and seven of cotton displayed low temperature-induced virescence when grown in controlled environments. For the group of plants studied, an increase in leaf chlorophyll a/b ratio was correlated with a temperature-sensitive biosynthetic sequence leading to a reduction in total chlorophyll content. These pigment alterations were reflected in the composition and quantity of the two major chlorophyll-protein complexes of chloroplast membranes. Changes in the amount of the major light-harvesting chlorophyll-protein complex was a prime consequence of the nuclear mutations. A decrease in the light-harvesting chlorophyll component of the light reaction centers of the leaf may account for the decrease in size of the photo-synthetic unit frequently noted in chlorophyll-deficient mutants. Variations in the concentration of the chlorophyll-protein complexes in the chloroplast lamellae may be causally related to variations in CO(2) compensation points of mutant soybean and cotton plants.

High photosynthetic rate of a chlorophyll mutant of cotton.

In a chlorophyll mutant (virescent) and wild-type cotton (Gossypium hirsutum L.), a number of photosynthetic parameters have been measured and compared with those published for other chlorophyll mutants. (a) The photosynthetic rates at 230 w/m(2) (400-700 nm) from a tungsten lamp were 36.8 mg CO(2) fixed/dm(2).hr (virescent) and 39.5 mg CO(2) fixed/dm(2).hr (wild-type). On a chlorphyll basis, the photosynthetic rates were 36.8 and 12.1 mg CO(2) fixed/mg chl.hr, respectively. (b) The photosynthetic rates at 13 w/m(2) (400-700 nm) from a tungsten source were 7.1 mg CO(2) fixed/dm(2).hr (virescent) and 7.4 mg CO(2) fixed/dm(2).hr (wild-type). On a chlorophyll basis, the photosynthetic rates were 6.0 and 1.4 mg CO(2) fixed/mg chl.hr, respectively. (c) The chlorophyll a/b ratios of the virescent and wild-type leaves were 3.3 and 4.1 (d) The chlorophyll/carotenoid ratios for the virescent and wild-type leaves were 3.2 and 7.3, respectively. (e) The photosynthetic carbon metabolism of the chlorophyll mutant was through the reductive pentose phosphate cycle. (f) The CO(2) compensation points for the virescent and wild-type plants were similar. (g) The mutant and wild-type leaves have the same quantum yield in the red part of the visible spectrum, but the virescent leaves have a lower quantum yield in the blue part of the spectrum. (h) Virescent and wild-type leaves contain similar levels on a protein basis of several reductive pentose phosphate cycle enzymes.

Characteristics of a virescent cotton mutant.

The virescent cotton (Gossypium hirsutum) mutant described here differs from normal cultivated cotton by a single mutation in the nucleus. The mutant exhibits nuclear control of chlorophyll and carotenoid development. Young leaves are distinctly yellow and become green with age. There is no unusual photometabolism of (14)CO(2) or (14)C-acetate in this mutant. It is probable that the nuclear virescent mutation is in a locus concerned with making structural units. The yellow leaves do show a high photosynthetic capacity on a chlorophyll basis. At saturating light intensity the rate of CO(2) fixation is 8 fold higher than the green control leaves. Thus, impaired pigment synthesis which could be lethal is offset by a high photosynthetic capacity in the virescent leaves.