Naturally colored cotton for wearable applications.

Naturally colored cotton (NCC) offers an environmentally friendly fiber for textile applications. Processing white cotton fiber into textiles requires extensive energy, water, and chemicals, whereas processing of NCC skips the most polluting activity, scouring-bleaching and dyeing; therefore, NCC provides an avenue to minimize the harmful impacts of textile production. NCC varieties are suitable for organic agriculture since they are naturally insect and disease-resistant, salt and drought-tolerant. Various fiber shades, ranging from light green to tan and brown, are available in the cultivated NCC (Gossypium hirsutum L.) species. The pigments responsible for the color of brown cotton fiber are proanthocyanidins or their derivatives synthesized by the flavonoid pathway. Due to pigments, the NCC has excellent ultraviolet protection properties. Some brown cotton varieties exhibited superior thermal resistance of fiber that can be used to make fabrics with enhanced flame retardancy. Here, we review molecular mechanisms involved in the pigment production of brown cotton and challenges in breeding NCC varieties with a wide range of colors but without penalty in fiber quality. Also, we discuss opportunities for NCC with flame-retarding properties in textile applications.

Multi-omics analysis of pigmentation related to proanthocyanidin biosynthesis in brown cotton (Gossypium hirsutum L.).

Naturally-colored brown cotton (NBC) fiber is an environmentally friendly raw source of fiber for textile applications. The fiber of some NBC cultivars exhibits flame-retardant properties, which can be used in textiles that require flame resistance. Proanthocyanidins or their derivatives are responsible for the brown pigment in NBC; however, how flame retardancy is related to pigmentation in NBC is poorly understood. To gain insight into brown pigment biosynthesis, we conducted comparative transcripts and metabolites profiling analysis of developing cotton fibers between the brown (MC-BL) and white (MC-WL) cotton near-isogenic lines (NILs), genetically different only in the Lc1 locus. In this study, mass spectrometry was used to detect metabolites in BL and WL developing fibers at 8, 12, 16, 20, 24, 36, and 40 days post anthesis (DPA) and mature fibers. Transcripts analysis was performed at two critical fiber developmental points, 8 DPA (fiber elongation) and 20 DPA (secondary cell wall deposition). We found 5836 (ESI MS positive mode) and 4541 (ESI MS negative mode) metabolites significantly different accumulated between BL and WL. Among them, 142 were known non-redundant metabolites, including organic acids, amino acids, and derivatives of the phenylpropanoid pathway. Transcript analysis determined 1691 (8 DPA) and 5073 (20 DPA) differentially expressed genes (DEGs) between BL and WL, with the majority of DEGs down-regulated at 20 DPA. Organic acids of the citric acid cycle were induced, while most of the detected amino acids were reduced in the MC-BL line. Both cis- and trans-stereoisomers of flavan-3-ols were detected in developing MC-WL and MC-BL fibers; however, the gallocatechin and catechin accumulated multiple times higher. Gas chromatography-mass spectrometry (GC-MS) analysis of fatty acids determined that palmitic acid long-chain alcohols were the main constituents of waxes of mature fibers. Energy-dispersive X-ray spectrometry (EDS) analysis of mature fibers revealed that potassium accumulated three times greater in MC-BL than in MC-WL mature fibers. This study provides novel insights into the biosynthesis of pigments and its association with flame retardancy in NBC fibers.

Preparation and Activity of Hemostatic and Antibacterial Dressings with Greige Cotton/Zeolite Formularies Having Silver and Ascorbic Acid Finishes.

The need for prehospital hemostatic dressings that exert an antibacterial effect is of interest for prolonged field care. Here, we consider a series of antibacterial and zeolite formulary treatment approaches applied to a cotton-based dressing. The design of the fabric formulations was based on the hemostatic dressing TACGauze with zeolite Y incorporated as a procoagulant with calcium and pectin to facilitate fiber adherence utilizing silver nanoparticles, and cellulose-crosslinked ascorbic acid to confer antibacterial activity. Infra-red spectra were employed to characterize the chemical modifications on the dressings. Contact angle measurements were employed to document the surface hydrophobicity of the cotton fabric which plays a role in the contact activation of the coagulation cascade. Ammonium Y zeolite-treated dressings initiated fibrin equal to the accepted standard hemorrhage control dressing and showed similar improvement with antibacterial finishes. The antibacterial activity of cotton-based technology utilizing both citrate-linked ascorbate-cellulose conjugate analogs and silver nanoparticle-embedded cotton fibers was observed against Staphylococcus aureus and Klebsiella pneumoniae at a level of 99.99 percent in the AATCC 100 assay. The hydrogen peroxide levels of the ascorbic acid-based fabrics, measured over a time period from zero up to forty-eight hours, were in line with the antibacterial activities.

Washable Antimicrobial Wipes Fabricated from a Blend of Nanocomposite Raw Cotton Fiber.

In this study, a simple and effective way to produce washable antimicrobial wipes was developed based on the unique ability of raw cotton fiber to produce silver nanoparticles. A nanocomposite substructure of silver nanoparticles (25 ± 3 nm) was generated in raw cotton fiber without reducing and stabilizing agents. This nanocomposite raw cotton fiber (2100 ± 58 mg/kg in the concentration of silver) was blended in the fabrication of nonwoven wipes. Blending small amounts in the wipes-0.5% for antimicrobial properties and 1% for wipe efficacy-reduced the viability of S. aureus and P. aeruginosa by 99.9%. The wipes, fabricated from a blend of 2% nanocomposite raw cotton fiber, maintained their antibacterial activities after 30 simulated laundering cycles. The washed wipes exhibited bacterial reductions greater than 98% for both Gram-positive and Gram-negative bacteria.

Flame resistant cotton lines generated by synergistic epistasis in a MAGIC population.

Textiles made from cotton fibers are flammable and thus often include flame retardant additives for consumer safety. Transgressive segregation in multi-parent populations facilitates new combinations of alleles of genes and can result in traits that are superior to those of any of the parents. A screen of 257 recombinant inbred lines from a multi-parent advanced generation intercross (MAGIC) population for naturally enhance flame retardance (FR) was conducted. All eleven parents, like all conventional white fiber cotton cultivars produce flammable fabric. MAGIC recombinant inbred lines (RILs) that produced fibers with significantly lower heat release capacities (HRC) as measured by microscale combustion calorimetry (MCC) were identified and the stability of the phenotypes of the outliers were confirmed when the RILs were grown at an additional location. Of the textiles fabricated from the five superior RILs, four exhibited the novel characteristic of inherent flame resistance. When exposed to open flame by standard 45° incline flammability testing, these four fabrics self-extinguished. To determine the genetic architecture of this novel trait, linkage, epistatic and multi-locus genome wide association studies (GWAS) were conducted with 473k SNPs identified by whole genome sequencing (WGS). Transcriptomes of developing fiber cells from select RILs were sequenced (RNAseq). Together, these data provide insight into the genetic mechanism of the unexpected emergence of flame-resistant cotton by transgressive segregation in a breeding program. The incorporation of this trait into global cotton germplasm by breeding has the potential to greatly reduce the costs and impacts of flame-retardant chemicals.

Thermal properties and surface chemistry of cotton varieties mineralized with calcium carbonate polymorphs by cyclic dipping.

In this study, hydroentangled cotton nonwovens were identified as effective hosts for mineralization of calcium carbonate (CaCO3) polymorphs to modify and improve their properties. All cotton varieties studied, including raw white cotton, scoured white cotton, and raw brown cotton, readily crystallized CaCO3 via a simple cyclic dipping process. A combination of analyses agreed that the surface chemistry of cotton fibers influenced the formation of different CaCO3 polymorphs. Scoured white cotton that consisted of almost pure cellulose predominantly produced the most stable calcite, whereas raw white and raw brown cottons that contain proteins facilitated the production of partial metastable vaterite. The morphology of calcite was better defined on the scoured cotton. The mineralization altered the hydrophobic surface of raw cottons to be hydrophilic, i.e., two-fold increase in moisture regain and decrease in water contact angle from 130 to 0 degrees. The mineralized cottons also exhibited improved thermal resistance, i.e., slower thermal decomposition with decreased activation energies and reduction in heat release capacity by up to 40%.

Role of xyloglucan in cotton (Gossypium hirsutum L.) fiber elongation of the short fiber mutant Ligon lintless-2 (Li2).

Xyloglucan is a matrix polysaccharide found in the cell walls of all land plants. In growing cells, xyloglucan is thought to connect cellulose microfibrils and regulate their separation during wall extension. Ligon lintless-2 (Li2) is a monogenic dominant cotton fiber mutation that causes extreme reduction in lint fiber length with no pleiotropic effects on vegetative growth. Li2 represents an excellent model system to study fiber elongation. To understand the role of xyloglucan in cotton fiber elongation we used the short fiber mutant Li2 and its near isogenic wild type for analysis of xyloglucan content and expression of xyloglucan-related genes in developing fibers. Accumulation of xyloglucan was significantly higher in Li2 developing fibers than in wild type. Genes encoding enzymes for nine family members of xyloglucan biosynthesis were identified in the draft Gossypium hirsutum genome. RNAseq analysis revealed that most differentially expressed xyloglucan-related genes were down-regulated in Li2 fiber cells. RT-qPCR analysis revealed that the peak of expression for the majority of xyloglucan-related genes in wild type developing fibers was 5-16days post anthesis (DPA) compared to 1-3 DPA in Li2 fibers. Thus, our results suggest that early activation of xyloglucan-related genes and down regulation of xyloglucan degradation genes during the elongation phase lead to elevated accumulation of xyloglucan that restricts elongation of fiber cells in Li2.

RNA-Seq Analysis of Developing Pecan (Carya illinoinensis) Embryos Reveals Parallel Expression Patterns among Allergen and Lipid Metabolism Genes.

The pecan nut is a nutrient-rich part of a healthy diet full of beneficial fatty acids and antioxidants, but can also cause allergic reactions in people suffering from food allergy to the nuts. The transcriptome of a developing pecan nut was characterized to identify the gene expression occurring during the process of nut development and to highlight those genes involved in fatty acid metabolism and those that commonly act as food allergens. Pecan samples were collected at several time points during the embryo development process including the water, gel, dough, and mature nut stages. Library preparation and sequencing were performed using Illumina-based mRNA HiSeq with RNA from four time points during the growing season during August and September 2012. Sequence analysis with Trinotate software following the Trinity protocol identified 133,000 unigenes with 52,267 named transcripts and 45,882 annotated genes. A total of 27,312 genes were defined by GO annotation. Gene expression clustering analysis identified 12 different gene expression profiles, each containing a number of genes. Three pecan seed storage proteins that commonly act as allergens, Car i 1, Car i 2, and Car i 4, were significantly up-regulated during the time course. Up-regulated fatty acid metabolism genes that were identified included acyl-[ACP] desaturase and omega-6 desaturase genes involved in oleic and linoleic acid metabolism. Notably, a few of the up-regulated acyl-[ACP] desaturase and omega-6 desaturase genes that were identified have expression patterns similar to the allergen genes based upon gene expression clustering and qPCR analysis. These findings suggest the possibility of coordinated accumulation of lipids and allergens during pecan nut embryogenesis.

A MAGIC population-based genome-wide association study reveals functional association of GhRBB1_A07 gene with superior fiber quality in cotton.

BACKGROUND: Cotton supplies a great majority of natural fiber for the global textile industry. The negative correlation between yield and fiber quality has hindered breeders' ability to improve these traits simultaneously. A multi-parent advanced generation inter-cross (MAGIC) population developed through random-mating of multiple diverse parents has the ability to break this negative correlation. Genotyping-by-sequencing (GBS) is a method that can rapidly identify and genotype a large number of single nucleotide polymorphisms (SNP). Genotyping a MAGIC population using GBS technologies will enable us to identify marker-trait associations with high resolution. RESULTS: An Upland cotton MAGIC population was developed through random-mating of 11 diverse cultivars for five generations. In this study, fiber quality data obtained from four environments and 6071 SNP markers generated via GBS and 223 microsatellite markers of 547 recombinant inbred lines (RILs) of the MAGIC population were used to conduct a genome wide association study (GWAS). By employing a mixed linear model, GWAS enabled us to identify markers significantly associated with fiber quantitative trait loci (QTL). We identified and validated one QTL cluster associated with four fiber quality traits [short fiber content (SFC), strength (STR), length (UHM) and uniformity (UI)] on chromosome A07. We further identified candidate genes related to fiber quality attributes in this region. Gene expression and amino acid substitution analysis suggested that a regeneration of bulb biogenesis 1 (GhRBB1_A07) gene is a candidate for superior fiber quality in Upland cotton. The DNA marker CFBid0004 designed from an 18 bp deletion in the coding sequence of GhRBB1_A07 in Acala Ultima is associated with the improved fiber quality in the MAGIC RILs and 105 additional commercial Upland cotton cultivars. CONCLUSION: Using GBS and a MAGIC population enabled more precise fiber QTL mapping in Upland cotton. The fiber QTL and associated markers identified in this study can be used to improve fiber quality through marker assisted selection or genomic selection in a cotton breeding program. Target manipulation of the GhRBB1_A07 gene through biotechnology or gene editing may potentially improve cotton fiber quality.

The GhTT2_A07 gene is linked to the brown colour and natural flame retardancy phenotypes of Lc1 cotton (Gossypium hirsutum L.) fibres.

Some naturally coloured brown cotton fibres from accessions of Gossypium hirsutum L. can be used to make textiles with enhanced flame retardancy (FR). Several independent brown fibre loci have been identified and mapped to chromosomes, but the underlying genes have not yet been identified, and the mechanism of lint fibre FR is not yet fully understood. In this study, we show that both the brown colour and enhanced FR of the Lc1 lint colour locus are linked to a 1.4Mb inversion on chromosome A07 that is immediately upstream of a gene with similarity to Arabidopsis TRANSPARENT TESTA 2 (TT2). As a result of the alternative upstream sequence, the transcription factor GhTT2_A07 is highly up-regulated in developing fibres. In turn, genes in the phenylpropanoid metabolic pathway are activated, leading to biosynthesis of proanthocyanidins and accumulation of inorganic elements. We show that enhanced FR and anthocyanin precursors appear in developing brown fibres well before the brown colour is detectible, demonstrating for the first time that the polymerized proanthocyanidins that constitute the brown colour are not the source of enhanced FR. Identifying the particular colourless metabolite that provides Lc1 cotton with enhanced FR could help minimize the use of synthetic chemical flame retardant additives in textiles.

Small RNA sequencing and degradome analysis of developing fibers of short fiber mutants Ligon-lintles-1 (Li 1 ) and -2 (Li 2 ) revealed a role for miRNAs and their targets in cotton fiber elongation.

BACKGROUND: The length of cotton fiber is an important agronomic trait that directly affects the quality of yarn and fabric. Understanding the molecular basis of fiber elongation would provide a means for improvement of fiber length. Ligon-lintless-1 (Li 1 ) and -2 (Li 2 ) are monogenic and dominant mutations that result in an extreme reduction in the length of lint fiber on mature seeds. In a near-isogenic state with wild type cotton these two short fiber mutants provide an effective model system to study the mechanisms of fiber elongation. Plant miRNAs regulate many aspects of growth and development. However, the mechanism underlying the miRNA-mediated regulation of fiber development is largely unknown. RESULTS: Small RNA libraries constructed from developing fiber cells of the short fiber mutants Li 1 and Li 2 and their near-isogenic wild type lines were sequenced. We identified 24 conservative and 147 novel miRNA families with targets that were detected through degradome sequencing. The distribution of the target genes into functional categories revealed the largest set of genes were transcription factors. Expression profiles of 20 miRNAs were examined across a fiber developmental time course in wild type and short fiber mutations. We conducted correlation analysis between miRNA transcript abundance and the length of fiber for 11 diverse Upland cotton lines. The expression patterns of 4 miRNAs revealed significant negative correlation with fiber lengths of 11 cotton lines. CONCLUSIONS: Our results suggested that the mutations have changed the regulation of miRNAs expression during fiber development. Further investigations of differentially expressed miRNAs in the Li 1 and Li 2 mutants will contribute to better understanding of the regulatory mechanisms of cotton fiber development. Four miRNAs negatively correlated with fiber length are good candidates for further investigations of miRNA regulation of important genotype dependent fiber traits. Thus, our results will contribute to further studies on the role of miRNAs in cotton fiber development and will provide a tool for fiber improvement through molecular breeding.

Phytohormonal networks promote differentiation of fiber initials on pre-anthesis cotton ovules grown in vitro and in planta.

The number of cotton (Gossypium sp.) ovule epidermal cells differentiating into fiber initials is an important factor affecting cotton yield and fiber quality. Despite extensive efforts in determining the molecular mechanisms regulating fiber initial differentiation, only a few genes responsible for fiber initial differentiation have been discovered. To identify putative genes directly involved in the fiber initiation process, we used a cotton ovule culture technique that controls the timing of fiber initial differentiation by exogenous phytohormone application in combination with comparative expression analyses between wild type and three fiberless mutants. The addition of exogenous auxin and gibberellins to pre-anthesis wild type ovules that did not have visible fiber initials increased the expression of genes affecting auxin, ethylene, ABA and jasmonic acid signaling pathways within 1 h after treatment. Most transcripts expressed differentially by the phytohormone treatment in vitro were also differentially expressed in the ovules of wild type and fiberless mutants that were grown in planta. In addition to MYB25-like, a gene that was previously shown to be associated with the differentiation of fiber initials, several other differentially expressed genes, including auxin/indole-3-acetic acid (AUX/IAA) involved in auxin signaling, ACC oxidase involved in ethylene biosynthesis, and abscisic acid (ABA) 8'-hydroxylase an enzyme that controls the rate of ABA catabolism, were co-regulated in the pre-anthesis ovules of both wild type and fiberless mutants. These results support the hypothesis that phytohormonal signaling networks regulate the temporal expression of genes responsible for differentiation of cotton fiber initials in vitro and in planta.

The Li2 mutation results in reduced subgenome expression bias in elongating fibers of allotetraploid cotton (Gossypium hirsutum L.).

Next generation sequencing (RNA-seq) technology was used to evaluate the effects of the Ligon lintless-2 (Li2) short fiber mutation on transcriptomes of both subgenomes of allotetraploid cotton (Gossypium hirsutum L.) as compared to its near-isogenic wild type. Sequencing was performed on 4 libraries from developing fibers of Li2 mutant and wild type near-isogenic lines at the peak of elongation followed by mapping and PolyCat categorization of RNA-seq data to the reference D5 genome (G. raimondii) for homeologous gene expression analysis. The majority of homeologous genes, 83.6% according to the reference genome, were expressed during fiber elongation. Our results revealed: 1) approximately two times more genes were induced in the AT subgenome comparing to the DT subgenome in wild type and mutant fiber; 2) the subgenome expression bias was significantly reduced in the Li2 fiber transcriptome; 3) Li2 had a significantly greater effect on the DT than on the AT subgenome. Transcriptional regulators and cell wall homeologous genes significantly affected by the Li2 mutation were reviewed in detail. This is the first report to explore the effects of a single mutation on homeologous gene expression in allotetraploid cotton. These results provide deeper insights into the evolution of allotetraploid cotton gene expression and cotton fiber development.

A transcript profiling approach reveals an abscisic acid-specific glycosyltransferase (UGT73C14) induced in developing fiber of Ligon lintless-2 mutant of cotton (Gossypium hirsutum L.).

Ligon lintless-2, a monogenic dominant cotton (Gossypium hirsutum L.) fiber mutation, causing extreme reduction in lint fiber length with no pleiotropic effects on vegetative growth, represents an excellent model system to study fiber elongation. A UDP-glycosyltransferase that was highly expressed in developing fibers of the mutant Ligon lintless-2 was isolated. The predicted amino acid sequence showed ~53% similarity with Arabidopsis UGT73C sub-family members and the UDP-glycosyltransferase was designated as UGT73C14. When expressed in Escherichia coli as a recombinant protein with a maltose binding protein tag, UGT73C14 displayed enzymatic activity toward ABA and utilized UDP-glucose and UDP-galactose as the sugar donors. The recombinant UGT73C14 converted natural occurring isoform (+)-cis, trans-ABA better than (+)-trans, trans-ABA and (-)-cis, trans-ABA. Transgenic Arabidopsis plants constitutively overexpressing UGT73C14 did not show phenotypic changes under standard growth conditions. However, the increased glycosylation of ABA resulted in phenotypic changes in post-germinative growth and seedling establishment, confirming in vivo activity of UGT73C14 for ABA. This suggests that the expression level of UGT73C14 is regulated by the observed elevated levels of ABA in developing fibers of the Li 2 mutant line and may be involved in the regulation of ABA homeostasis.

Integrated metabolomics and genomics analysis provides new insights into the fiber elongation process in Ligon lintless-2 mutant cotton (Gossypium hirsutum L.).

BACKGROUND: The length of cotton fiber is an important agronomic trait characteristic that directly affects the quality of yarn and fabric. The cotton (Gossypium hirsutum L.) fiber mutation, Ligon lintless-2, is controlled by a single dominant gene (Li(2)) and results in extremely shortened lint fibers on mature seeds with no visible pleiotropic effects on vegetative growth and development. The Li(2) mutant phenotype provides an ideal model system to study fiber elongation. To understand metabolic processes involved in cotton fiber elongation, changes in metabolites and transcripts in the Li(2) mutant fibers were compared to wild-type fibers during development. RESULTS: Principal component analysis of metabolites from GC-MS data separated Li(2) mutant fiber samples from WT fiber samples at the WT elongation stage, indicating that the Li(2) mutation altered the metabolome of the mutant fibers. The observed alterations in the Li(2) metabolome included significant reductions in the levels of detected free sugars, sugar alcohols, sugar acids, and sugar phosphates. Biological processes associated with carbohydrate biosynthesis, cell wall loosening, and cytoskeleton were also down-regulated in Li(2) fibers. Gamma-aminobutyric acid, known as a signaling factor in many organisms, was significantly elevated in mutant fibers. Higher accumulation of 2-ketoglutarate, succinate, and malate suggested higher nitrate assimilation in the Li(2) line. Transcriptional activation of genes involved in nitrogen compound metabolism along with changes in the levels of nitrogen transport amino acids suggested re-direction of carbon flow into nitrogen metabolism in Li(2) mutant fibers. CONCLUSIONS: This report provides the first comprehensive analysis of metabolite and transcript changes in response to the Li(2) mutation in elongating fibers. A number of factors associated with cell elongation found in this study will facilitate further research in understanding metabolic processes of cotton fiber elongation.

Cloning and characterization of homeologous cellulose synthase catalytic subunit 2 genes from allotetraploid cotton (Gossypium hirsutum L.).

Cellulose synthase catalytic subunits (CesAs) are the catalytic sites within a multisubunit complex for cellulose biosynthesis in plants. CesAs have been extensively studied in diploid plants, but are not well characterized in polyploid plants. Gossypium hirsutum is an allotetraploid cotton species producing over 90% of the world's cotton fibers. Although G. hirsutum CesAs (GhCesAs) are responsible for cellulose production in cotton fiber, very limited numbers of GhCesA genes have been identified. Here, we report isolating and characterizing a pair of homeologous CesA2 genes and their full-length cDNAs from allotetraploid cotton. The GhCesA2-A(T) gene from the A-subgenome and GhCesA2-D(T) gene from the D-subgenome were screened from a G. hirsutum BAC library. These genes shared 92% sequence similarity throughout the entire sequence. The coding sequences were nearly identical, and the deduced amino acid sequences from GhCesA2-A(T) (1,039 amino acids) and GhCesA2-D(T) (1,040 amino acids) were identical except four amino acids, whereas the noncoding sequences showed divergence. Sequence analyses showed that all exons of GhCesA2-A(T) contained consensus splice donor dinucleotides, but one exon in GhCesA2-D(T) contained nonconsensus splice donor dinucleotides. Although the nonconsensus splice donor dinucleotides were previously suggested to be involved in alternative splice or pseudogenization, our results showed that a majority of GhCesA2-A(T) and GhCesA2-D(T) transcripts consisted of functional and full-length transcripts with little evidence for alternative mRNA isoforms in developing cotton fibers. Expression analyses showed that GhCesA2-A(T) and GhCesA2-D(T) shared common temporal and spatial expression patterns, and they were highly and preferentially expressed during the cellulose biosynthesis stage in developing cotton fibers. The observations of higher expression levels of both GhCesA2-A(T) and GhCesA2-D(T) in developing fibers of one near-isogenic line (NIL) with higher fiber bundle strength over the other NIL with lower fiber bundle strength suggested that the differential expression of genes associated with secondary cell wall cellulose biosynthesis in developing fiber might affect cotton fiber properties.

A combined functional and structural genomics approach identified an EST-SSR marker with complete linkage to the Ligon lintless-2 genetic locus in cotton (Gossypium hirsutum L.).

BACKGROUND: Cotton fiber length is an important quality attribute to the textile industry and longer fibers can be more efficiently spun into yarns to produce superior fabrics. There is typically a negative correlation between yield and fiber quality traits such as length. An understanding of the regulatory mechanisms controlling fiber length can potentially provide a valuable tool for cotton breeders to improve fiber length while maintaining high yields. The cotton (Gossypium hirsutum L.) fiber mutation Ligon lintless-2 is controlled by a single dominant gene (Li2) that results in significantly shorter fibers than a wild-type. In a near-isogenic state with a wild-type cotton line, Li2 is a model system with which to study fiber elongation. RESULTS: Two near-isogenic lines of Ligon lintless-2 (Li2) cotton, one mutant and one wild-type, were developed through five generations of backcrosses (BC5). An F2 population was developed from a cross between the two Li2 near-isogenic lines and used to develop a linkage map of the Li2 locus on chromosome 18. Five simple sequence repeat (SSR) markers were closely mapped around the Li2 locus region with two of the markers flanking the Li2 locus at 0.87 and 0.52 centimorgan. No apparent differences in fiber initiation and early fiber elongation were observed between the mutant ovules and the wild-type ones. Gene expression profiling using microarrays suggested roles of reactive oxygen species (ROS) homeostasis and cytokinin regulation in the Li2 mutant phenotype. Microarray gene expression data led to successful identification of an EST-SSR marker (NAU3991) that displayed complete linkage to the Li2 locus. CONCLUSIONS: In the field of cotton genomics, we report the first successful conversion of gene expression data into an SSR marker that is associated with a genomic region harboring a gene responsible for a fiber trait. The EST-derived SSR marker NAU3991 displayed complete linkage to the Li2 locus on chromosome 18 and resided in a gene with similarity to a putative plectin-related protein. The complete linkage suggests that this expressed sequence may be the Li2 gene.

Near-isogenic cotton germplasm lines that differ in fiber-bundle strength have temporal differences in fiber gene expression patterns as revealed by comparative high-throughput profiling.

Gene expression profiles of developing cotton (Gossypium hirsutum L.) fibers from two near-isogenic lines (NILs) that differ in fiber-bundle strength, short-fiber content, and in fewer than two genetic loci were compared using an oligonucleotide microarray. Fiber gene expression was compared at five time points spanning fiber elongation and secondary cell wall (SCW) biosynthesis. Fiber samples were collected from field plots in a randomized, complete block design, with three spatially distinct biological replications for each NIL at each time point. Microarray hybridizations were performed in a loop experimental design that allowed comparisons of fiber gene expression profiles as a function of time between the two NILs. Overall, developmental expression patterns revealed by the microarray experiment agreed with previously reported cotton fiber gene expression patterns for specific genes. Additionally, genes expressed coordinately with the onset of SCW biosynthesis in cotton fiber correlated with gene expression patterns of other SCW-producing plant tissues. Functional classification and enrichment analysis of differentially expressed genes between the two NILs revealed that genes associated with SCW biosynthesis were significantly up-regulated in fibers of the high-fiber quality line at the transition stage of cotton fiber development. For independent corroboration of the microarray results, 15 genes were selected for quantitative reverse transcription PCR analysis of fiber gene expression. These analyses, conducted over multiple field years, confirmed the temporal difference in fiber gene expression between the two NILs. We hypothesize that the loci conferring temporal differences in fiber gene expression between the NILs are important regulatory sequences that offer the potential for more targeted manipulation of cotton fiber quality.

Resistance gene analogue markers are mapped to homeologous chromosomes in cultivated tetraploid cotton.

Degenerate primers designed from conserved motifs of known plant resistance gene products were used to amplify genomic DNA sequences from the root-knot nematode (Meloidogyne incognita) resistance genetic source, Upland cotton (Gossypium hirsutum) cultivar Auburn 634 RNR. A total of 165 clones were isolated, and sequence analysis revealed 57 of the clones to be novel nucleotide sequences, many containing the resistance (R)-protein nucleotide-binding site motif. A cluster analysis was performed with resistance gene analogue (RGA) nucleotide sequences isolated in this study, in addition to 99 cotton RGA nucleotide sequences already deposited in GenBank, to generate a phylogenetic tree of cotton R genes. The cotton RGA nucleotide sequences were arranged into 11 groups and 56 sub-groups, based on genetic distances. Multiple sequence alignments were performed on the RGA sequences of each sub-group, and either the consensus sequences or individual RGA sequences were used to design 61 RGA-sequence-tagged site primers. A recombinant inbred line (RIL) population of cultivated tetraploid cotton was genotyped using RGA-specific primers that amplified polymorphic fragments between the two RIL parents. Nine RGA markers were mapped to homeologous chromosomes 12 and 26, based on linkage to existing markers that are located on these chromosomes.