Comparative chromosome painting between two marsupials: origins of an XX/XY1Y2 sex chromosome system

Cross-species chromosome painting was used to investigate genome rearrangements between tammar wallaby Macropus eugenii (2n = 16) and the swamp wallaby Wallabia bicolor (2n = 10♀/11♂), which diverged about 6 million years ago. The swamp wallaby has an XX female:XY₁Y₂ male sex chromosome system thought to have resulted from a fusion between an autosome and the small original X, not involving the Y. Thus, the small Y₁ should represent the original Y and the large Y₂ the original autosome. DNA paints were prepared from flow-sorted and microdissected chromosomes from the tammar wallaby. Painting swamp wallaby spreads with each tammar chromosome-specific probe gave extremely strong and clear signals in single-, two-, and three-color FISH. These showed that two tammar wallaby autosomes are represented unchanged in the swamp wallaby, two are represented by different centric fusions, and one by a tandem fusion to make the very long arms of swamp wallaby Chromosome (Chr) 1. The large swamp wallaby X comprises the tammar X as its short arm, and a tandemly fused 7 and 2 as the long arm. The acrocentric swamp wallaby Y₂ is a 2/7 fusion, homologous with the long arm of the X. The small swamp wallaby Y₁ is confirmed as the original Y by its painting with the tammar Y. However, the presence of sequences shared between the microdissected tammar Xp and Y on the swamp wallaby Y₂ implies that the formation of the compound sex chromosomes involved addition of autosome(s) to both the original X and Y. We propose that this involved fusion with an ancient pseudoautosomal region followed by fission proximal to this shared region. Received: 16 October 1996/Accepted: 30 January 1997     

Plant regeneration from mesophyll protoplasts of Matthiola incana (L.) R. Br.

Summary A protocol for obtaining regenerated fertile plants from mesophyll protoplasts of three lines of Matthiola incana is described. Protoplasts were isolated from leaves of 21–28 days old Matthiola plants grown in controlled environment. Sustained divisions were achieved when protoplasts were embedded in sodium alginate. Up to 2.0 % of the protoplasts developed into colonies which could be transferred to shoot regeneration media. More than 25 % of the obtained calluses regenerated shoots. About 4 % of these shoots could be rooted and after transfer to soil phenotypically normal plants have been obtained.Abbreviations 2,4-D 2,4-dichlorphenoxyacetic acid - NAA naphthalene acetic acid - IAA indole-3-acetic acid - BAP 6-banzylaminopurine - IPA isopentenyladenine - IPAR isopentenyladenosine - MES (2-[N-morpholino]) ethanesulfonic acid     

RS46(DXS548) genotyping of reproductive cells: approaching preimplantation testing of the fragile-X syndrome

In order to approach preimplantation testing for the fragile-X syndrome, we used genotyping of the polymorphic RS46(DXS548) locus closely linked to the FMR1 gene, in single reproductive cells of females. The RS46(DXS548) amplification was adjusted to the single cell level by a two-round polymerase chain reaction (PCR) procedure. Unfertilized oocytes and extruded polar bodies were subjected to PCR. RS46(DXS548) genotyping at the single cell level was successful in 95% of the samples. In two-third of the metaphase II oocytes and first polar bodies obtained from women who were heterozygous at the RS46(DXS548) locus, both maternal RS46(DXS548) alleles were observed because of crossing over during the first meiotic division. This makes gamete selection by first polar body analysis inefficient. From the allele frequencies found in 56 unrelated individuals, a heterozygote frequency of 51% was estimated, whereas the observed heterozygote frequency was 56%. The whole PCR procedure can be performed within 16 h after blastomere biopsy. Consequently, the selection and transfer of the diagnosed embryos can be carried out within an acceptable time. Therefore, preimplantation testing for the fragile-X syndrome with the RS46(DXS548) AC-repeat may be an alternative choice for prenatal testing for those carrier females who are heterozygous (informative) at the RS46(DXS548) locus.     

Reproductive strategies of coastal marine fishes in the tropics

Synopsis A synthesis of ethnobiological, behavioral and physical oceanographic information leads to the conclusion that temperate zone models of reproductive strategy are inapplicable to many fishes of the coastal tropics. Intense predation appears to exert heavy selection pressure on fishes that spend their adult lives in coral, mangrove or tropical seagrass communities. Many exhibit spawning behaviors and spawn at times and locations that favor the transport of their pelagic eggs and pelagic larvae offshore where predation is reduced. This creates a countervailing selection pressure — the need to return the larvae to shallow water once they are ready to colonize their post-larval habitats. Accordingly, spawning is often concentrated at times of the year when prevailing winds or currents are at their weakest, thereby reducing the transport of larvae long distances from where they originated. Spawning is also concentrated in the vicinity of nearshore gyres which similarly favor the ultimate return of the larvae to their natal area. Among these species, therefore, offshore larval dispersal does not seem to be an adaptation for dispersal of the species, but rather an evolutionary response to intense predation pressure in the adult habitats. Lunar reproductive periodicity is more common among these species than has previously been recognized, and is one of the strategies employed to enhance the offshore flushing of eggs and larvae.This paper forms part of the proceedings of a mini-symposium convened at Cornell University, Ithaca, N. Y. 18–19 May 1976, entitled Patterns of Community Structure in Fishes (G. S. Helfman, ed.).Contribution No. 524, Hawaii Institute of Marine Biology, University of Hawaii, Honolulu.     

Conversion of Biovolume Measurements of Soil Organisms, Grown Under Various Moisture Tensions, to Biomass and Their Nutrient Content

Direct microscopic measurements of biomass in soil require conversion factors for calculation of the mass of microorganisms from the measured volumes. These factors were determined for two bacteria, five fungi, and a yeast isolated from soil. Moisture stress conditions occurring in nature were simulated by growth in two media using shake cultures, on agar plates, and on membranes held at 34, 330, and 1,390 kPa of suction. The observed conversion factors, i.e., the ratio between dry weight and wet volume, generally increased with increasing moisture stress. The ratios for fungi ranged from 0.11 to 0.41 g/cm³ with an average of 0.33 g/cm³. Correction of earlier data assuming 80% water and a wet-weight specific gravity of 1.1 would require a conversion factor of 1.44. The dry-weight specific gravity of bacteria and yeasts ranged from 0.38 to 1.4 g/cm³ with an average of 0.8 g/cm³. These high values can only occur at 10% ash if no more than 50% of the cell is water, and a specific conversion factor to correct past data for bacterial biomass has not yet been suggested. The high conversion factors for bacteria and yeast could not be explained by shrinkage of cells due to heat fixing, but shrinkage during preparation could not be completely discounted. Moisture stress affected the C, N, and P content of the various organisms, with the ash contents increasing with increasing moisture stress. Although further work is necessary to obtain accurate conversion factors between biovolume and biomass, especially for bacteria, this study clearly indicates that existing data on the specific gravity and the water and nutrient content of microorganisms grown in shake cultures cannot be applied when quantifying the soil microbial biomass.     

Energization of the sugar transport mechanism in the plasmalemma of isolated mesophyll protoplasts

The mechanism of 3-O-methyl-d-glucose transport through the plasmalemma has been investigated in protoplasts isolated from the mesophyll of Pisum sativum L. var. Dan.Analysis of the fluxes after 50 minutes of uptake showed that the gradual decrease in slope of the net uptake curve with time was not due to any decline in uptake capacity; it represented the approach to flux equilibrium of a small compartment of the protoplast, probably the cytoplasm.The energy of activation for initial flux into this compartment was 20 kilocalories per mole between 17 and 27 C. Very high discrimination was shown with regard to sugar isomers. Light strongly promoted flux (by a factor of 2.5 in the case of methyl glucose). Initial flux showed sharply contrasting inhibitor sensitivity in the light and the dark. Light uptake was sensitive to the proton conductor carbonyl cyanide m-chlorophenylhydrazone (CCCP), but stable for at least the first 10 minutes to the ATPase inhibitors quercetin, rutin, and diethylstilbestrol, as well as to arsenate. Dark uptake, on the other hand, was stable to CCCP but was immediately depressed by quercetin, rutin, diethylstilbestrol, and arsenate.Protoplasts which received a light pretreatment before incubation in the dark took up methyl glucose at the accelerated light rate for the first 7 minutes. Moreover, the light pretreatment sensitized subsequent initial dark uptake to CCCP, and conferred on it the stability to ATPase inhibitors and arsenate characteristic of light uptake. After about 7 minutes the characteristic inhibitor responses of dark uptake were resumed.It is proposed that more than one mode of energy-coupling for sugar transport may operate in these protoplasts.     

Hysteresis in the responses of membrane potential, membrane resistance, and growth rate to cyclic temperature change

Measurements of electrical potential, membrane resistance, and elongation rate have been carried out on the developing pollen tube of Oenothera drummondii.     

Ectothiorhodospira vacuolata sp. nov., a new phototrophic bacterium from soda lakes

A new phototrophic bacterium was isolated from Jordanian and Kenyan alkaline salt lakes. Cells are rod shaped, 1.5 m wide and 2–4 m long, and motile by polar flagella. They divide by binary fission, and possess photosynthetic membranes as lamellar stacks similar to those in the other species of the genus Ectothiorhodospira and the brown colored Rhodospirillum species. The presence of bacteriochlorophyll a and carotenoids of the normal spirilloxanthin series is indicated by the absorption spectra of living cells. Under certain growth conditions the cells form gas vacuoles, may become immotile and float to the top of the culture medium. Sulfide and thiosulfate are used as photosynthetic electron donors. During the oxidation of sulfide to sulfate, elemental sulfur is formed, which is accumulated outside the cells. The organisms are strictly anaerobic, do not require vitamins, are moderately halophilic and need alkaline pH-values for growth. The new species Ectothiorhodospira vacuolata is proposed.     

Determination of the configurations of lactic and glyceric acids from human serum and urine by capillary gas—liquid chromatography

The separation of the enantiomers of lactic and glyceric acids can be achieved by capillary gas chromatography on SP-1000 using the corresponding O-acetylated menthyl esters. The structures of the derivatives were proved by proton magnetic resonance spectroscopy and mass spectrometry. The method has been used for the determination of the absolute configurations of lactic and glyceric acids isolated from serum and urine from different patients.