Transfer of defined numbers of chloroplasts into albino protoplasts using an improved subprotoplast/protoplast microfusion procedure: Transfer of only two chloroplasts leads to variegated progeny

Summary A procedure is described by which it is possible to perform controlled microfusion of microscopically selected protoplast fusion partners with high efficiencies. The procedure is applied to fusion of Nicotiana tabacum (line 92V37, N. undulata cytoplasm) plastid albino protoplasts as a recipient and spontaneously formed subprotoplasts of green N. tabacum (line SRI) as donor. Products of individual electrofusion events are cloned via single cell nurse culture and the derived cell lines are analysed for the occurrence of variegated or green regenerating shoots, which are indicative of the establishment of the transferred organelles in the cell progeny. The plastid population in green regenerants recovered after the transfer of only two chloroplasts was demonstrated to have originated from the donor subprotoplast organelles by restriction analysis of total DNA using a plastome-specific probe.Some of the results described in this paper have been presented as posters at scientific meetings (Eigel and Koop 1989b; Eigel and Koop 1990)     

Windkanalversuche zur Abh?ngigkeit der Fluggeschwindigkeit einer Wei?scheitelammer (Zonotrichia leucophrys) von der Windgeschwindigkeit

Zusammenfassung In einem Windkanal wurde für eine Weißscheitelammer (Zonotrichia leucophrys) die Abhängigkeit der Fluggeschwindigkeit in Luft (v_F) und der Geschwindigkeit über Grund (v_G) von der Windgeschwindigkeit (v_W) bestimmt. Das Versuchstier flog bei Windstille mit einer mittleren v_F von 5,15 m/sec. Bei zunehmenden Gegenwinden wurde v_F so stark erhöht, daß sich v_G nur langsam verringerte. Bei v_W=16,2 m/sec wurde noch v_G=1,4 m/sec erreicht. Bei zunehmenden Rückenwinden wurde v_F immer mehr herabgesetzt, so daß sich v_G kaum erhöhte. Bei v_W=–7 m/sec flog das Versuchstier sogar langsam rückwärts in der Luft, um seine Manövrierfähigkeit im engen Kanal zu erhalten. Der Gesamtgeschwindigkeitsbereich für v_F lag zwischen –0,9 bis 17,6 m/sec.

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Flight speed of a White-Crowned Sparrow in a Wind Tunnel at different Wind Speeds

Summary In an experimental wind tunnel air speed (v_F) and ground speed (v_G) of a White-Crowned Sparrow (Zonotrichia leucophrys) were measured as a function of wind speed (v_W). The bird had an average v_F of 5,15 m/sec in still air. With rising head winds it increased its v_F. At v_W=16,2 m/sec v_F was 17,6 m/sec and v_G was still 1,4 m/sec. With tailwinds it tended to maintain constant v_G and lowered v_F. At v_W=–7 m/sec it even flew slowly backward in the air in order to maintain its maneuvrability. The complete speed range for v_F was from –0,9 to + 17,6 m/sec.

     

Industrial asymmetric catalysis: Approaches and results

In this short overview problems, requirements and strategies for the application of enantioselective catalysis in fine chemicals production are discussed. The factors are described and analyzed which determine whether a catalytic method is suitable for industrial application. Selected examples of enantioselective catalytic production processes are described in some detail in order to illustrate the potential of this modern technology.

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Riassunto   Catalisi asimmetrica in dustriale: Metodiche e risultati. In questa breve rassegna vengono discussi i requisiti e le strategie applicative per l'impiego della catalisi, enantioselettiva nella produzione di fine chemicals. Sono descritti ed analizzati i fattori che determinano l'applicabilità industriale di un metodo catalitico. Vengono scelti e descritti in qualche dettaglio esempi di processi produttivi catalitici enantioselettivi, che servono ad illustrare il potenziale della tecnologia moderna.

     

A highly enantioselective rhodium catalyst for the hydrogenation of aliphatic α-keto esters

The enantioselective hydrogenation of several α-keto acid derivatives with rhodium diphosphine catalysts has been investigated using a random screening approach. The neutral rhodium catalyst prepared in situ from bis(2,5-norbornadiene rhodium chloride) and NORPHOS has been found to be an excellent catalyst for preparing aliphatic α-hydroxy esters in high optical purities. The reaction parameters for the hydrogenation of ethyl 2-oxo-4-phenyl-butyrate, an intermediate for the ACE inhibitor Benazepril, were optimized and the best optical yields obtained were 96%.     

The complete amino acid sequence of hirudin,a thrombin specific inhibitor: Application of colour carboxymethylation

Color carboxymethylation of cysteine residues with a new chromophoric reagent dimethylaminoazobenzene iodoacetamide, was applied to the micro-sequence analysis of hirudin, a thrombin specific inhibitor. Six cysteine residues of the reduced hirudin were detected as colored phenylthiohydantoin derivative and 3 tryptic peptides of hirudin (all containing cysteines) were isolated as colored peptide. The complete hirudin sequence, including 6 uncertain positions left in the previous report [Petersen T.E. (1976) in: Protides of the Biological Fluids; 23rd Colloquium, pp. 145, Pergamon Press, London] was established.     

Effects of site specific amino acid exchanges in hirudin on the thrombin-hirudin interaction

For the identification of the primary binding site of hirudin for thrombin we generated hirudin mutants with site directed amino acid substitutions with the help of recombinant DNA technology. Preliminary results indicate, that lys (47) may be directly involved in the hirudin-thrombin interaction: 1. The mutant glu (47) shows a Ki-value which is increased by two orders of magnitude (1.6.10(-9) M); 2. Incubation of mutant ala(48) with endoproteinase lys-C results in proteolysis of the newly formed peptide bond lys(47)-ala(48), whereas all other peptide bonds (lys-X) are not accessible.     

The problem of 4,5-dioxovaleric acid as a precursor of 5-aminolevulinic acid and chlorophyll in green algae

During chlorophyll biosynthesis, 4,5-dioxovaleric acid is supposed to be an intermediate between 2-oxoglutarate and 5-aminolevulinic acid. Although the occurrence of 4,5-dioxovaleric acid in cells and culture filtrates of unicellular green algae treated with levulinic acid has been previously reported, improvement of the analytical method now shows that 4,5-dioxovaleric acid does not occur in these organisms but arises due to interference of 5-aminolevulinic acid in 4,5-dioxovaleric acid analysis. The significance of 4,5-dioxovaleric acid as a free metabolite in 5-aminolevulinic acid biosynthesis must therefore be reconsidered.     

Die Ultraschall-Ortungslaute der Hufeisen-Fledermäuse (Chiroptera-Rhinolophidae) in verschiedenen Orientierungssituationen

Zusammenfassung Rhinolophus ferrum-equinum (RF) und Rhinolophus euryale (RE) erzeugen in allen Orientierungssituationen dreiteilige Ortungslaute. Im nur wenig intensiven Anfangsteil steigt die Frequenz um 1–12 KHz bis zur Frequenz des folgenden Mittelteils an. Der Mittelteil nimmt mindestens 9/10 der Laute ein und hat eine konstante Frequenz von etwa 83 KHz bei RF und 104 KHz bei RE. Im Endteil fällt die Frequenz immer um 13–16 KHz ab. Entsprechend dem Atemrhythmus werden die Ortungslaute in Folgen von langen Einzellauten oder in Lautgruppen ausgesendet. Sie sind in der Vertikalen und der Horizontalen gleich stark gebündelt. 20,5° seitlich von der Vorausrichtung fällt die Intensität auf die Hälfte der Vorausintensität ab.In Situationen, in denen die Fledermäuse nicht unmittelbar ein Objekt anpeilen, wie beim ungestörten Hängen, beim Start und beim freien Flug, senden sie mit einer Wiederholfrequenz von 4–10 Hz lange Einzellaute aus, deren Dauer meist zwischen 50–65 msec bei RF und 35–45 msec bei RE liegt. Diese Laute haben ein ungefähr in der Lautmitte liegendes Intensitätsmaximum. Beim gezielten Anpeilen eines Objekts, wie beim Peilen nach einem vorgehaltenen Mehlwurm, beim Flug durch Hindernisse und bei der Landung, werden die Laute verkürzt bis zu minimal 10 msec bei RF und 7 msec bei RE und in Gruppen ausgesendet. Die Wiederholfrequenz steigt an bis zu maximal 70–80 Hz bei RF und 100 Hz bei RE und im frequenzmodulierten Endteil ergibt sich ein zweites Intensitätsmaximum. Bei der Landung wird die Dauer der Ortungslaute und ebenso die Dauer des frequenzmodulierten Endteiles in linearer Abhängigkeit von der Entfernung zur Landestange verkürzt.Fliegende Tiere senken die Frequenz des konstantfrequenten Mittelteils immer um den Betrag der durch die Fluggeschwindigkeit bedingten Dopplereffekte ab, so daß die von den Tieren gehörte Frequenz nahezu konstant in Höhe der vor dem Flug ausgesendeten Frequenz gehalten wird. Eine hängende und auf ein sich bewegendes Pendel peilende RF war ebenfalls bestrebt, die durch die Pendelbewegung entstehenden Dopplereffekte zu kompensieren und die Frequenz der gehörten Echos konstant zu halten.Beide Arten zeigen beim Flug durch Hindernisse ähnliche Ortungsleistungen. Sie sind bei horizontalen und vertikalen Drahthindernissen noch in der Lage Drähte von 0,08 mm Durchmesser zu vermeiden. Die untere Ortungsgrenze liegt bei RF zwischen 0,08-0,05 mm und bei RE etwa bei 0,05 mm. Der mittlere Abstand, bei dem Fledermäuse auf ein Hindernis reagieren, fällt von 140 cm bei 3 mm Drahtdurchmesser auf 38 cm bei 0,08 mm ab. Bei 0,05 mm reagierte RE in 3 von 8 Flügen noch auf das Hindernis, während RF keine Reaktion mehr zeigte.

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The ultrasonic sounds of horseshoe bats (Chiroptera-Rhinolophidae) in different orientation situations

Summary Echolocating Rhinolophus ferrum-equinum (RF) and Rhinolophus euryale (RE) produce ultrasonic pulses which always consist of three parts. In the initial part the frequency rises by 1–12 KHz reaching finally a frequency of about 83 KHz in RF and about 104 KHz in RE. These frequencies remain constant throughout the middle part which comprises at least 9/10 of the pulses. The terminal part is frequency-modulated. The frequency drops by 13–16 KHz. The animals emit either long pulses or groups of short pulses corresponding to the respiratory cycle.In situations, in which the bats are not echolocating a special object, e.g. while hanging, taking off or free flying, they emit long pulses with a repetition rate of 4–10 Hz and a duration of 50–65 msec in RF and 35–45 msec in RE. These pulses have their maximum intensity in the middle. When echolocating directly to an object, e.g. while echolocating a mealworm, passing an obstacle or landing, the pulses are emitted in groups and shortened down to a minimum duration of 10 msec in RF and 7 msec in RE, while the repetition rate rises to 702–80 Hz in RF and 100 Hz in RE. At the same time a second intensity maximum appears in the frequency-modulated terminal part. While landing, the pulses as well as the terminal parts are shortened in a linear relation to the distance from the landing bar.Flying animals lower the frequency of the middle part by such an amount that the Doppler shifts caused by the flight velocity are compensated. Therefore the frequency heard by the bats remains constant and is as high as the frequency emitted before the flight. One hanging RF even tried to compensate the Doppler shifts caused by a moving pendulum in order to keep the echo frequency constant.While passing obstacles animals of both species show a similar orientation performance. They are able to avoid wires down to a wire diameter of 0.08 mm. The minimum limit for detection lies between 0.08–0.05 mm in RF and about 0.05 mm in RE. The mean distance at which the bats react to an obstacle descreases from 140 cm at a wire diameter of 3 mm to 38 cm at 0.08 mm. Even at 0.05 mm RE changed its sound sequence in 3 of 8 flights at a distance of 18 cm from the obstacle, whereas RF did not react any more.