Protein polymorphism and genetic divergence in slow loris (genusNycticebus)

In this study, protein electrophoresis was assayed to detect genetic variation in GenusNycticebus. A total of 29 samples (2N. coucang and 27N. pygmaeus) were analyzed for 42 genetic loci. In the 27 samples ofN. pygmaeus, 4 loci were observed to be polymerphic. Therefore, the estimatedP value (proportion of polymorphic loci) is 0.095, theA value (average number of alleles each locus) is 1.045, and theH value (mean individual heterozygosity) is 0.040. After comparing theH ofN. pygmaeus with those of other primates reported, we found that the protein variation inN. pygmaeus is slightly lower than the average level. Additionally, we also observed obivious allele difference betweenN. pygmaeus andN. coucang. There are no shared alleles between these two species in eight loci. TheNei's genetic distance between them was calculated as 0.2541, which falls in the spectrum of genetic difference between species in primates.     

Scanning electron microscopy of microspore embryogenesis inBrassica spp.

Scanning electron microscopy was employed to study and compare microspore embryogenesis in vitro with pollen development in planta inBrassica napus andB. oleracea. An exine with its specific pattern had already been formed, when microspores were released from tetrads. During subsequent pollen development, microspores increased in size and continued to strengthen the exine. Upon in vitro culture, all microspores, i.e., embryogenic and nonembryogenic, initially showed the same morphological features. After 24 h in culture, the microspores had increased in size. Thereafter, embryogenesis was indicated in some microspores by two different morphological changes. One featured an expansion in volume of the cell cluster around the germination aperture (type I), the other showed cell cluster volume expansion over the entire microspore surface (type II). Two-thirds of embryogenic microspores in bothB. napus andB. oleracea demonstrated type I development. When followed by fluorescence microscopy, in vitro culture of microspores revealed cultures with a high embryo frequency were those with a high frequency of symmetrical division.Abbreviations SEM Scanning electron microscopy - TEM Transmission electron microscopy     

Genetic divergence among some maintainer and restorer lines in relation to hybrid breeding in rice (Oryza sativa L.)

Summary A representative group of 100 elite lines, 67 of which are restorers and 33 maintainers, from 68 crosses made at IRRI and 18 improved varieties from five countries were studied, using Mahalonobis, D²-statistic and canonical analysis, to understand the nature and magnitude of divergence and to assess the importance of a set of quantitative characters related to yield in genetic differentiation. The 100 genotypes were grouped into 13 clusters. There were three single variety clusters and the number of lines in the remaining clusters ranged from 2–36. Canonical vector values indicated the importance of yield followed by 1,000-grain weight, days to maturity and plant height in primary as well as in secondary differentiation. Results showed that yield, number of tillers per plant, days to maturity and 1,000-grain weight contributed largely to the divergence. There were no indications of a relationship between geographical diversity and genetic diversity in the present study. Disposition of IRRI developed maintainers and restorers into various clusters indicated the presence of large amounts of diversity within the IRRI elite lines which suggested that these materials could be used in crossing programs to produce heterotic F₁ hybrids. Crossing of maintainers and restorers among the highly diverse groups was suggested as it may produce F₁S that will give higher magnitudes of heterosis. Part of M.S. Thesis of the senior author     

Genetic divergence among thirty-one genotypes of toria (Brassica campestris L.) grown in twelve environments

Summary Thirty-one toria genotypes were grown in 12 artificially created environments in order to study genetic divergence. D² estimates based on 12 characters were used in obtaining the clustering pattern and inter- and intracluster distances. Selection of divergent parents has been made in three ways, i.e., on the basis of (a) genetic divergence exhibited when grown in the richest and most productive environment; (b) stable and consistent values of divergence over all 12 environments and (c) pooled analysis. Out of 31 genotypes, on the basis of stability, high yield and divergence six genotypes were found to be suitable for use in a breeding programme.     

In vitro versus in vivo genetic divergence in potato

 The objective of this study was to compare the genetic divergence pattern in potato (Solanum tuberosum L.) under in vitro and in vivo conditions. Twenty two potato genotypes were evaluated for ten morphological characters under four in vitro conditions, and for 20 characters under four in vivo seasons. Mahalanobis’ generalized intra- and inter-group genetic distances, and the distribution of genotypes into different clusters, led to the same conclusions under both in vitro and in vivo conditions: (1) genetic diversity was not related to geographic diversity, (2) genetic distances were higher between Tuberosum and Andigena than within Tuberosum and Andigena, and (3) present-day Indian varieties have more resemblance to Tuberosum than to the Andigena group. The in vitro approach was more effective than the in vivo approach for differentiating the genotypes per se, although its effectiveness for cross prediction is known to be low. Received: 15 September 1997 / Accepted: 15 July 1998     

Somatic embryogenesis and in vitro flowering inBrassica nigra

Summary This report describes a protocol for regeneration ofBrassica nigra in vitro from unorganized callus to a highly differentiated stage of flowering. Callus is initiated from seedling hypocotyl, and root explants and plantlets are obtained via somatic embryogenesis. Shoot cultures can be established from these plantlets. These shoots can either be induced to flower in vitro or rooted to produce plants which flower ex vitro. Each stage of development is marked with a specific growth regulator requirement. This has potential as a model system to understand the cellular and molecular mechanisms involved in morphogenesis, and it can be used to understand the mechanism of change of phase from vegetative to reproductive. An advantage of this system is that in vitro flowering can be obtained repeatedly in the shoots raised from the axillary buds of the flowering shoots. The protocol can also be used to procureB. nigra gametes under aseptic condition.     

Development of microsporesin vivo andin vitro inBrassica napus L.

Summary Populations of highly homogeneous uninucleate and binucleate microspores ofBrassica napus cv. Topas were obtained by bud selection and percoll fractionation. The development of the uninucleate and the binucleate microspores in culture was compared to thosein vivo using the fluorochrome DAPI to stain DNA. The major developmental pathway of the uninucleate microsporesin vitro resulted in embryo formation. The characteristic of this pathway was that the first division produced two diffusely stained nuclei and subsequent divisions gave rise to a multinucleate embryoid. The second pathway which occurred in a small number of the uninucleate microspores led to callus formation. The majority of the binucleate microsporesin vitro followed the developmental pattern of their counterpartsin vivo and were not embryogenic. The embryogenic binucleate microspores produced embryos through the divisions of the vegetative nucleus.Plant Research Centre Contribution # 1147     

Protein polymorphism inEligmodontia typus. Genetic divergence with other phyllotine cricetids

By means of gel electrophoresis of tissue extracts we have studied protein polymorphism inEligmodontia typus. The analysis was performed on specimens from five population samples collected at different sites in Patagonia (Argentina). Mean heterozygosity (\-h) and proportion of polymorphic loci (P) were determined on the basis of 19 loci. Considering all individuals as one sample, \-h gave a value of 0.16 and P was 70%. Although these values are much higher than those reported for most rodent species, they are very similar to those obtained by us for four species of the genusCalomys and forGraomys griseoflavus. There is a striking genetic identity (I_N=0.99) among populations from regions with different environmental conditions, indicating that the species possesses a common genic pool. Genetic distance with other species of the Phyllotini was estimated. D_N was lower betweenE. typus andCalomys (mean D_N=0.88) than betweenE. typus andGraomys griseoflavus (D_N=1.01). The high morphological similarity between these last two species, especially regarding those characters related to desert life adaptation, could be assigned, at least in part, to convergent evolution.     

Genetic divergence between two forms of a tongue sole <Emphasis Type="Italic">Cynoglossus interruptus</Emphasis>

Genetic differences between trilinear and dilinear forms of a tongue sole, Cynoglossus interruptus, were examined by use of isozymes. Unequivocal differences were detected between the two forms, including complete replacement of alleles at the FBALD-2*, MDH-3*, PROT-1*, and SOD-1* loci, almost complete replacement at the AAT-3*, AH-1*, AH-2*, EST-3*, G3PDH-3*, GPI-2*, IDHP-1*, and MDH-1* loci, and extreme differences in allelic frequencies at the GPI-1* and PGDH* loci. The genetic distances (D values) between the two forms were 0.4207–0.4353, figures predicatively significant at the specific level. The considerable genetic differences strongly suggested that the two forms represent distinct species. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Genetic divergence and phylogenetic analysis of genus <Emphasis Type="Italic">Jatropha</Emphasis> based on nuclear ribosomal DNA ITS sequence

The genus Jatropha belongs to the family Euphorbiaceae having significant economic importance. The present investigation was undertaken with an aim to understand phylogenetic relationships among seven species (J. curcas, J. glandulifera, J. gossypifolia, J. integerrima, J. multifida, J. podagrica, and J. tanjorensis.) which are widely distributed in India, using nuclear ribosomal DNA ITS sequence (nrDNA ITS) and to compare the results with multilocus marker analysis systems reported earlier for the same genus. The size variation obtained among sequenced nrDNA ITS regions was narrow and ranged from 647 to 654 bp. The overall mean genetic distance (GD) of genus Jatropha was found to be 0.385. Highest interspecific GD (0.419) was found between J. glandulifera and J. multifida. The least interspecific GD (0.085) was found between J. gossypifolia and J. tanjorensis. The highest intraspecific GD was observed in J. podagrica (0.011) and least in J. gossypifolia (0.002). The phylogram obtained using nrDNA ITS sequence showed congruence with the phylograms obtained using multilocus markers system reported earlier with minor variations. The present study also strongly supports high phylogentic closeness of J. curcas and J. integerrima. The only exception found was J. podagrica which clustered with J. multifida in earlier based on multilocus marker analysis, was clustered with J. curcas in the present analysis. The sequence data generated in the present investigation will help for further studies in intraspecies population, and their phylogentic analysis, biogeographical, molecular evolution studies and also pave way for future phylogetic and/or evolution studies among the other groups belongs to the family Euphorbiaceae.     

Mitochondrial DNA divergence in yoshinobori gobies (Rhinogobius species complex) between the Bonin Islands and the Japan–Ryukyu Archipelago

To investigate the genetic differentiation between the Bonin (Ogasawara) Islands' freshwater goby Rhinogobius sp. Bonin Island (BI) form (Ogasawara-yoshinobori) and the Japan–Ryukyu Archipelago relatives, the mitochondrial DNA (mtDNA) phylogeny of Japanese Rhinogobius species was inferred from partial nucleotide sequences of the mitochondrial NADH dehydrogenase 5 subunit (ND5) gene (945 bp). The resultant tree showed that the Bonin Islands group separated first from the other Japanese lineage, and a test calculation indicated the divergence date to be approximately 3 million years BP. Although it is necessary to use a more reliable estimate to confirm the divergence date, Rhinogobius sp. BI has retained its mtDNA lineage in the islands for millions of years.     

The genetic basis of adaptation to selection for longevity inDrosophila melanogaster

Summary Analysis of electrophoretic loci shows that at least four differences exist in isozymes of long- and short-lived populations ofD. melanogaster, descended by selection from a common ancestral stock. Adults of longlived populations differ in gene dosage of phosphoglucomutase (PGM), NAD malate dehydrogenase (MHD), NADP malic enzyme (ME) and by additional mobility variants of glucose-6-phosphate dehydrogenase (G6PD). Larvae, however, differ only by variants of G6PD. The differences in these enzymes, considered together with the greater flight endurance that long-lived populations have shown elsewhere, suggest that increased glycogen synthesis plays a significant role in the improved life span of selected populations. Adaptation to selection for increased life span may, therefore, derive from an improved ability to use dietary sucrose in the media provided. The distribution of electrophoretic loci agrees with the results of a study indicating the position of genetic elements contributing to life span.     

Direct regeneration of transformed shoots inBrassica napus from hypocotyl infections withAgrobacterium rhizogenes

Genetically transformed root clones of rapeseed (Brassica napus) were obtained afterin vitro infection of excised hypocotyl segments with a wild type strain ofAgrobacterium rhizogenes and two strains ofA. rhizogenes harbouring kanamycin resistance. The ability of hairy root formation was affected by light and was highly dependent on the location of the infection site at the hypocotyl. Inoculation of decapitated hypocotyls with an intact root system gave rise to direct shoot formation from the site of inoculation. Histological sections showed that several meristems were initiated at the inoculation site. Root and shoot clones were isolated and subcultured axenically in hormone-free liquid MS medium. Identification of transformed root and shoot clones was based on opine assays. Further selection was carried out in kanamycin-enriched medium.All opine-positive root clones showed NPT II (neomycin phosphotransferase) activity. Nearly half of the shoot clones expressed a strong NPT II activity while the rest gave a weak or no NPT II response.     

Utilization of wild relatives in genetic improvement of Arachis hypogaea L.

Summary The chromosome complements of 12 taxa in section Arachis were karyotypically and meiotically analysed. In taxa with 2n=20 the arm ratio of the respective pair of chromosomes was taken as an independent quantitative character and statistically analysed by Mahalanobis D². Two clusters were formed, one represented solely by A. batizocoi and the other consisting of the remaining 11 taxa. This grouping was confirmed by canonical analysis. In the larger group of species, A villosa and A. correntina were closely related karyotypically and on D² distance, while A. cardenasii forms a distinct subgroup. A. cardenasii lacks the short A chromosome recorded in other species of this group, and A. batizocoi is no longer the only species to have a pair of chromosomes with a secondary constriction. The taxa with 2n=40, A. monticola and A. hypogaea, are karyotypically very similar, though there is a difference in the number of chromosome pairs with a secondary constriction. On the basis of karyomorphological affinity, especially in relation to marker chromosomes, A. cardenasii is probably one of the ancestors of the tetraploid species studied.Approved as ICRISAT Journal Article No. 169 and released for publication     

Differential expression of members of the napin storage protein gene family during embryogenesis inBrassica napus

S1 nuclease analysis and sub-family-specific oligonucleotide probes were used to characterize the expression during embryogenesis of the napin storage protein gene family ofBrassica napus (oilseed rape). The expression of one sub-class represented by the napin gene gNa peaks and declines earlier than the other members of the family. This sub-class was highly expressed representing ca. 20% of napin mRNA at 26 days after anthesis.     

Interaction of cycloheximide and light on chlorophyll content and chloroplast ultrastructure inBrassica campestris

Cycloheximide retarded the loss of chlorophyll from detached komatsuna (Brassica campestris cv. Komatsuna) leaves during incubation in the dark but promoted its loss in light. Cycloheximide-induced chlorophyll bleaching in light was prevented by some active oxygen scavengers. Chloroplast envelopes of cycloheximide-treated leaves incubated in both the dark and light were destroyed within 48 h. The grana of cycloheximide-treated leaves incubated in light were dilated.     

Remarkable genetic divergence of Gymnodiptychus dybowskii between south and north of Tianshan Mountain in northwest China

Gymnodiptychus dybowskii is endemic to Xinjiang, China and has been locally listed as protected animals. To investigate its genetic diversity and structure, specimens were collected from six localities in Yili River system and Kaidu River. Fragments of 1092bp Cyt b gene were sequenced for 116 individuals. A total of 21 haplotypes were found in all samples, and no haplotype was shared between Yili River system and Kaidu River population. Sequence comparisons revealed 123 variable sites, with eight singleton sites and 115 parsimony informative sites. For all the populations examined, the haplotype diversity (h) was 0.8298 ± 0.0226, nucleotide diversity (π) was 0.2521 ± 0.1202, and average number of pairwise nucleotide differences (k) was 275.3369 ± 118.5660. AMOVA analysis showed that the differences were significant for total populations except for Yili River system populations. The pairwise F_st values revealed same conclusion with AMOVA analysis: Kaidu River population was divergent from Yili River system populations. The genetic distance between two groups was 0.108 and the divergence time was estimated at 5.4–6.6 Ma, the uplift of Tianshan Mountain might have separated them and resulted in the genetic differentiation. The neutrality test and mismatch analysis indicated that both two groups of G. dybowskii had went through population expansion, the expansion time of Yili River system and Kaidu River population was estimated at 0.5859–0.7146 Ma and 0.5151–0.6282 Ma, respectively. The climate changes of Qinghai-Tibetan Plateau might have influenced the demographic history of G. dybowskii.     

Stable sodium sulfate tolerance inBrassica napus cv. Westar callus cultures

Summary Sodium-sulfate-tolerant callus ofBrassica napus cv. Westar, selected on medium containing 105 mM Na₂SO₄, was maintained on medium without the salt to test for stability of tolerance. Tolerance to Na₂SO₄ was retained even after 18 subcultures on no salt. This tissue also showed tolerance to K₂SO₄, NaCl, and KCl. However, with the exception of callus grown on KCl fresh weight yields were less than that of tolerant callus maintained continuously on Na₂SO₄. Tolerant callus maintained on no salt had a mixture of the compact morphology of unselected callus and the friable morphology of tolerant callus. Both callus types expressed salt tolerance. Sucrose, reducing sugars and proline concentrations were measured in unselected callus, tolerant callus maintained continuously on Na₂SO₄, and tolerant callus maintained on no salt. Sucrose levels were similar in all cases. Whether maintained on or off Na₂SO₄., tolerant callus had reducing sugars levels three to fou times greater than unselected callus. Tolerant callus maintained on no salt had twice the amount of proline found in the unselected callus. Tolerant callus maintained in the absence of salt had an ash content, sodium concentration, and potassium concentration significantly lower than that of unselected callus. Supported by the Natural Sciences and Engineering Research Council of Canada Strategic Grant nos. G 0949 and G 1642 to T. A. Thorpe, D. M. Reid, and E.C. Yeung.     

Allozyme divergence among species ofWolffia (Lemnaceae)

The genusWolffia was surveyed electrophoretically at 14 allozyme loci. A total of 133 clones representing 10 of the 11 recognized species was examined. Genetic identities among most pairs of species are zero, with non-zero values ranging from 0.14 to 0.40.Wolffia angusta and the newly describedW. neglecta show the highest similarity, and the former species has an identity of 0.14 withW. australiana. The next highest similarity (0.34) occurs betweenW. globosa of Southeast Asia andW. cylindracea of southern Africa, which until recently, had generally been viewed as members of the same species. Other species showing some common alleles are members of a complex involvingW. arrhiza, W. columbiana, W. cylindracea, andW. globosa. WithinW. arrhiza, plants from South Africa and Europe are easily distinguished electrophoretically because each contains unique alleles at two loci. Strains from other parts of Africa vary at these loci and are not totally distinct from either the plants from South Africa or from Europe. Species ofWolffia are much more divergent at allozyme loci than the majority of congeners of flowering plants. This suggests that the species are quite old and that the difficulties in distinguishing taxa morphologically are the result of reduction rather than lack of divergence due to recent speciation. Because of the lack of shared alleles between the majority of species pairs inWolffia, enzyme electrophoresis provides limited resolution of species relationships in the genus.