Applications of genome mapping and marker-assisted selection (MAS) in crop improvement are reviewed. The following aspects are considered: a comparison of the choice of markers available for the generation of linkage maps (including amplified fragment length polymorphisms (AFLP); restriction fragment length polymorphisms (RFLP); randomly amplified polymorphic DNA (RAPD) and simple sequence repeats (SSR)); quantitative trait loci (QTL) analysis; use of molecular markers in the exploitation of hybrid vigour; physical genome mapping; map-based cloning and transposon tagging of agriculturally important genes; synteny in cereal genomes; and the use of MAS in breeding for disease and pest resistance.

Genome mapping, molecular markers and marker-assisted selection in crop plants

Calling female moths attract their mates late at night with intermittent release of a species-specific sex-pheromone blend Mean frequency of pheromone filaments encodes distance to the calling female In their zig-zagging upwind search male moths encounter turbulent pheromone blend filaments at highly variable concentrations and frequencies The male moth antennae are delicately designed to detect and distinguish even traces of these sex pheromones amongst the abundance of other odors Its olfactory receptor neurons sense even single pheromone molecules and track intermittent pheromone filaments of highly variable frequencies up to about 30 Hz over a wide concentration range In the hawkmoth Manduca sexta brief, weak pheromone stimuli as encountered during flight are detected via a metabotropic PLCβ-dependent signal transduction cascade which leads to transient changes in intracellular Ca2+ concentrations Strong or long pheromone stimuli, which are possibly perceived in direct contact with the female, activate receptor-guanylyl cyclases causing long-term adaptation In addition, depending on endogenous rhythms of the moth´s physiological state, hormones such as the stress hormone octopamine modulate second messenger levels in sensory neurons High octopamine levels during the activity phase maximize temporal resolution cAMP-dependently as a prerequisite to mate location Thus, I suggest that sliding adjustment of odor response threshold and kinetics is based upon relative concentration ratios of intracellular Ca2+ and cyclic nucleotide levels which gate different ion channels synergistically In addition, I propose a new hypothesis for the cyclic nucleotide-dependent ion channel formed by insect olfactory receptor/coreceptor complexes Instead of being employed for an ionotropic mechanism of odor detection it is proposed to control subthreshold membrane potential oscillation of sensory neurons, as a basis for temporal encoding of odors

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Pheromone Transduction in Moths

Bitter gourd (Momordica charantia) is an important vegetable and medicinal plant in tropical and subtropical regions globally. In this study, the draft genome sequence of a monoecious bitter gourd inbred line, OHB3-1, was analyzed. Through Illumina sequencing and de novo assembly, scaffolds of 285.5 Mb in length were generated, corresponding to ∼84% of the estimated genome size of bitter gourd (339 Mb). In this draft genome sequence, 45,859 protein-coding gene loci were identified, and transposable elements accounted for 15.3% of the whole genome. According to synteny mapping and phylogenetic analysis of conserved genes, bitter gourd was more related to watermelon (Citrullus lanatus) than to cucumber (Cucumis sativus) or melon (C. melo). Using RAD-seq analysis, 1507 marker loci were genotyped in an F2 progeny of two bitter gourd lines, resulting in an improved linkage map, comprising 11 linkage groups. By anchoring RAD tag markers, 255 scaffolds were assigned to the linkage map. Comparative analysis of genome sequences and predicted genes determined that putative trypsin-inhibitor and ribosome-inactivating genes were distinctive in the bitter gourd genome. These genes could characterize the bitter gourd as a medicinal plant.

/pdf/draft-genome-sequence-of-bitter-gourd-momordica-charantia-a-37p503xyrr.pdf

Draft genome sequence of bitter gourd (Momordica charantia), a vegetable and medicinal plant in tropical and subtropical regions

Insect olfactory receptors are heteromeric ligand-gated ion channels composed of at least one common subunit (Orco) and at least one subunit that confers odorant specificity. Little is known about how individual subunits contribute to the structure and function of the olfactory receptor complex. We expressed insect olfactory receptors in Xenopus oocytes to investigate 2 functional features, ion channel block and odorant recognition. The sensitivity of Drosophila olfactory receptors to inhibition by ruthenium red, a cation channel blocker, varied widely when different specificity subunits were present, suggesting that the specificity subunits contribute to the structure of the ion pore. Olfactory receptors formed by Dmel\Or35a and Orco subunits from several different species displayed highly similar odorant response profiles, suggesting that the Orco subunit does not contribute to the structure of the odorant-binding site. We further explored odorant recognition by conducting a detailed examination of the odorant specificity Dmel\Or67a + Dmel\Orco, a receptor that responds to aromatic structures. This screen identified agonists, partial agonists, and an antagonist of Dmel\Or67a + Dmel\Orco. Our findings favor specific subunit arrangements within the olfactory receptor complex and provide a preliminary odorophore for an olfactory receptor, offering a useful foundation for future exploration of insect olfactory receptor structure.

/pdf/subunit-contributions-to-insect-olfactory-receptor-function-16ymas8wk7.pdf

Subunit Contributions to Insect Olfactory Receptor Function: Channel Block and Odorant Recognition

Food prospects and nutraceutical attributes of Momordica species: A potential tropical bioresources – A review

Somatic chromosome number and detailed karyotype analysis were carried out in six Indian Momordica species viz. M. balsamina, M. charantia, M. cochinchinensis, M. dioica, M. sahyadrica and M. cymbalaria (syn. Luffa cymbalaria; a taxon of controversial taxonomic identity). The somatic chromosome number 2n = 22 was reconfirmed in monoecious species (M. balsamina and M. charantia). Out of four dioecious species, the chromosome number was reconfirmed in M. cochinchinensis (2n = 28), M. dioica (2n = 28) and M. subangulata subsp. renigera (2n = 56), while in M. sahyadrica (2n = 28) somatic chromosome number was reported for the first time. A new chromosome number of 2n = 18 was reported in M. cymbalaria against its previous reports of 2n = 16, 22. The karyotype analysis of all the species revealed significant numerical and structural variations of chromosomes. It was possible to distinguish chromosomes of M. cymbalaria from other Momordica species and also between monoecious and dioecious taxa of the genus. Morphology and crossability among the dioecious species was also studied. Evidence from morphology, crossability, pollen viability and chromosome synapsis suggests a segmental allopolyploid origin for M. subangulata subsp. renigera. The taxonomic status of the controversial taxon M. cymbalaria was also discussed using morphological, karyological and crossability data.

Cytotaxonomical analysis of Momordica L. (Cucurbitaceae) species of Indian occurrence.

The homologues of yeast INO80 are identified across phyla from Caenorhabditis elegans to human. In Drosophila it has been shown that dINO80 forms a complex with Pleiohomeotic but does not interact with Hox PRE (polycomb responsive element). As some proteins of the INO80 complex are implicated in homeotic gene regulation, we examined if dINO80 is involved in regulation of homeotic genes. We find that dINO80 null mutants generated by imprecise excision of P-element are late embryonic lethals and show homeotic transformation. We detect misexpression of homeotic genes like Sex-comb reduced, Antennapedia, Ultrabithorax and Abdominal-B in dIno80 mutant embryos by immunostaining which is further substantiated by quantitative PCR. Polycomb phenotype in dIno80-Pc is enhanced in double mutants. Concurrently, the localization of dINO80 to sequences upstream of misexpressed genes in vivo shows that dINO80 is involved in homeotic gene regulation and probably through its interactions with PcG-trxG complexes.

Chromatin remodeling protein INO80 has a role in regulation of homeotic gene expression in Drosophila.

In this paper, we show that mutants in the gene stambhA (stmA), which encodes a putative phosphatidylinositol 4,5 bisphosphate-diacylglycerol lipase, exhibit a significant reduction in the amplitudes of odor-evoked responses recorded from the antennal surface of adult Drosophila. This lends support to previously published findings that olfactory transduction in Drosophila requires a phospholipid intermediate. Mutations in stmA also affect the olfactory behavior response of larvae. Moreover, there is a requirement for Gqα and phospholipase Cβ function in larval olfaction. The results suggest that larval olfactory transduction, like that of the adult, utilizes a phospholipid second messenger, generated by the activation of Gqα and Plcβ21c, and modulated by the stmA gene product.

Drosophila mutants in phospholipid signaling have reduced olfactory responses as adults and larvae.

Random Amplified Polymorphic DNA (RAPD) analysis was used to evaluate genetic diversity among commercial Indian cotton varieties. Fifteen varieties belonging to Gossypium hirsutum L and seven to G. arboreum L were analyzed with 50 random decamer primers using the polymerase chain reaction (PCR). Twenty six of the primers detected polymorphism in all 22 cotton varieties. A total of 371 bands were amplified, 87% of which were polymorphic. Cluster analysis by the unweighted pair group method of arithmetic means (UPGMA) showed that diploids and tetraploids can be divided in two groups at a similarity of 30%. Diploid variety C402W showed the least similarity to all the others in the group. Among tetraploids, closely related varieties Pusa 8-6, 4515 and RS 875 were distinctly different from the rest. The analysis revealed that the intervarietal genetic relationships of several varieties is related to their pedigree. The results also revealed that tetraploids show a much narrower genetic base (similarity range of 65–95%) than the diploids (similarity range 54–88%). The results obtained can be used for the selection of parents to generate a mapping population and begin a breeding programme.

RAPD Analysis of Genetic Diversity in Indian Tetraploid and Diploid Cotton (Gossypium spp)

Studies on meiosis in pollen mother cells (PMCs) of a triploid interspecific hybrid (3x = 39 chromosomes, AAD) between tetraploid Gossypium hirsutum (4n = 2x = 52,AADD) and diploid G. arboreum (2n = 2x = 26,AA) are reported. During meiotic metaphase I, 13 AA bivalents and 13 D univalents are expected in the hybrid. However, only 28% of the PMCs had this expected configuration. The rest of the PMCs had between 8 and 12 bivalents and between 12 and 17 univalents. Univalents lagged at anaphase I, and at metaphase II one or a group of univalents remained scattered in the cytoplasm and failed to assemble at a single metaphase plate. Primary bipolar spindles organized around the bivalents and multivalents. In addition to the primary spindle, several secondary and smaller bipolar spindles organized themselves around individual univalents and groups of univalents. Almost all (97%) of the PMCs showed secondary spindles. Each spindle functioned independently and despite their multiple numbers in a cell, meiosis I proceeded normally, with polyad formation. These observations strongly support the view that in plant meiocytes bilateral kinetochore symmetry is not required for establishing a bipolar spindle and that single unpaired chromosomes can initiate and stabilize the formation of a functional bipolar spindle.

Shanti Chandrashekaran

Papers

Cytotaxonomical analysis of Momordica L. (Cucurbitaceae) species of Indian occurrence.

Chromatin remodeling protein INO80 has a role in regulation of homeotic gene expression in Drosophila.

Drosophila mutants in phospholipid signaling have reduced olfactory responses as adults and larvae.

RAPD Analysis of Genetic Diversity in Indian Tetraploid and Diploid Cotton (Gossypium spp)

Meiosis in a triploid hybrid of Gossypium: high frequency of secondary bipolar spindles at metaphase II.