Microalgae engineering toolbox: Selectable and screenable markers

Engineering microalgae has opened a new era for plant biologists and biotechnologists. Microalgae had been proved as a promising candidate for the production of biopharmaceuticals, nutraceuticals, antioxidants, antimicrobial and antiviral compounds, in dyeing and food industry as well for biofuel production. Genetic transformation of some important microalgae has been successful, but several other potential microalgae species still need scientific attention. The success of the genetic transformation depends mainly on the utilization of the selectable and screenable markers. Like for other higher crop plants, several useful markers have been reported for microalgae transformation. In this follow-up, we compared different marker genes for genetic engineering of approximately all the industrially important microalgae. We have discussed the expression host, the targeted genome, appropriate selection agent, as well as the transformation method. Genetic transformation is an expensive and labor intensive process and this review will aid to shorten the time span by providing a database of appropriate markers for microalgae research which could serve as a guide for those involved in the genetic engineering of microalgae.     

Release of the cellular prion protein from cultured cells after loss of its glycoinositol phospholipid anchor

Secreted forms of the sialoglycoprotein designated cellularprion protein (PrP^C) have been identified that cannot be explainedby alternative splicing. We report that secreted forms of PrP^Cderive from precursors that are bound to the plasma membraneby glycoinositol phospholipid (GPI) anchors. Secreted PrP^C slowlyappeared in the culture medium of metabolically radiolabelledcells after incubations of 8—24 h. Digestion of nascentPrP^C with phosphatidylinositol-specific phospholipase C (PIPLC)prevented the appearance of secreted PrP^C. Secreted PrP^c partitionedinto the aqueous phase of Triton X–114 like PIPLC-releasedPrP^C. While the M_r of PIPLC-released PrP^C was reduced 2–4kDa after treatment with aqueous hydroflouric acid, which removesthe entire GPI anchor modification, the M_r of secreted PrP^Cwas unchanged. Both PIPLC-released and secreted PrP^c were recognizedby antiserum raised against a synthetic C-terminal peptide correspondingto residues 220–233 (amino acid 231 is the site of GPIattachment). We conclude that GPI-anchored PrP^C is post-translationallyprocessed to remove most, if not all, of the GPI modificationand then shed into culture medium. Whether PrP^C is shed afterproteolysis near the C-terminus remains to be established. Aminority of PrP^C in normal Syrian hamster brain partitionedinto the aqueous phase of Triton X–114 like shed PrP^Csuggesting physiological significance. post-translational prion protein secretion sialoglycoprotein     

Prelaying exodus of Cory's Shearwaters (Calonectris diomedea borealis) on Selvagem Grande

Summary We stayed on the Portugese island of Selvagem Grande, 30° N 16° W, 300 km south of Madeira and 200 km north of Tenerife, from 9 May to 3 June 1991 in order to study the prelaying exodus in Cory's Shearwaters (Calonectris diomedea borealis) and its connection with the colony attendance cycles described earlier. Laying took place on 1.5 June ±3.4 days. During the preceding weeks males and females of Cory's Shearwater show very different attendance patterns. Females are absent for 19±4.8 days at minimum. Absences of males are much shorter and also much more variable. They last 7.9±6.0 days (coefficient of variation 76 %, as against 12 % in females). The first incubation stint of the males is badly synchronised with the laying: the delay between laying and the start of the males' first stint may amount up to one week, without compromising the breeding success. Two well marked attendance peaks have been observed, separated by an interval of 18 days, twice the length of that observed in June 1988. We suggest that this anomaly in the daily attendance graph is correlated with the prelaying exodus. The function of the prelaying exodus and its relation with the energy cost of producing the egg and the necessity of storing lipid reserves are discussed.

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Vorbrutzeitliches Verlassen der Kolonie beim Gelbschnabelsturmtaucher (Calonectris diomedea borealis) auf Selvagem Grande

Zusammenfassung Zwischen 9. Mai und 3. Juni 1991 untersuchten wir auf der portugisischen Insel Selvagem Grande, 30° N 16° W, 300 km südlich von Madeira und 200 km nördlich von Teneriffa gelegen, das Verlassen der Kolonie vor der Eiablage des Gelbschnabelsturmtauchers (Calonectris diomedea borealis) und die Beziehung zur Anwesenheit in der Kolonie. Gelegt wurde am 1,5 Juni ±3,4 Tage. In den Monaten vor der Eiablage zeigten Männchen und Weibchen sehr unterschiedliche Muster der Anwesenheit in der Kolonie. Während die Weibchen im Minimum durchschnittlich 19±4,8 Tage abwesend waren, war diese Abwesenheit bei den Männchen mit 7,9±6,0 Tagen wesentlich kürzer und variabler (Variationskoeffizient 76 % gegenüber 12 % bei den Weibchen). Die erste Brutbeteiligung des Männchens war kaum mit der Eiablage synchronisiert, doch war dadurch der Bruterfolg nicht beeinträchtigt. Die Anwesenheit in der Kolonie zeigte zwei Gipfel, die 18 Tage auseinander lagen. In 1988 betrug dieser zeitliche Abstand nur etwa die Hälfte. Es wird vermutet, dass dieser Unterschied mit dem Ausmaß des vorbrutzeitlichen Verlassens der Kolonie korreliert. Die Funktion dieses vorbrutzeitlichen Exodus und seine Beziehung zu den energetischen Kosten der Eiproduktion und der Bildung von Lipidreserven werden diskutiert.

     

Purification of the growth-related protein p25 of the Ehrlich ascites tumor and analysis of its isoforms

1. Two of the three isoforms of the growth-related protein p25 of the Ehrlich ascites tumor have been purified to homogeneity by giant two-dimensional polyacrylamide gel electrophoresis. 2. Antibodies raised against the isoform p25/1 react also with isoforms p25/2 and p25/3. 3. Limited tryptic digestion of p25/1 and p25/2 resulted in similar oligopeptide patterns. Corresponding oligopeptides of both isoforms have identical amino acid sequences. 4. The isoforms p25/2 and p25/3 are phosphorylated derivatives of unphosphorylated p25/1. The phosphorus is bound to serine and a further unknown phosphorylation site.     

Biochemical characterization of a putative axonal guidance molecule of the chick visual system

Temporal retinal axons growing in vitro on carpets of tectal membranes are deflected by cell membranes of posterior tectum. The activity responsible for this deflection can be abolished by antibodies raised against tectal membranes and the corresponding Fab fragments. Analysis of tectal membranes by two-dimensional gel electrophoresis and immunoblotting reveals a 33 kd glycoprotein that has a higher concentration in posterior than in anterior tectum. Its expression is developmentally regulated, and it is sensitive to phosphatidylinositol-specific phospholipase C. These are properties expected for a molecule responsible for the phenomena observed in experiments on in vitro guidance of retinal axons.     

The division between fast- and slow-growing species corresponds to natural relationships among the mycobacteria.

Comparative 16S rRNA sequencing was used to infer the phylogenetic relationships among selected species of mycobacteria and related organisms. The phylogeny inferred reflects the traditional classification, with major branches of the phylogenetic tree in general correspondence to the four Runyon groups and with numerical classification analyses. All the mycobacterial species compared, with the exception of M. chitae, are closely related (average similarity values greater than 95%). The slow growers form a coherent line of descent, distinct from the rapid growers, within which the overt pathogens are clustered. The distant relationship between M. chitae and the remaining mycobacteria suggests that this organism is incorrectly classified with the mycobacteria. M. paratuberculosis 18 was indistinguishable from M. avium-M. intracellulare-M. scrofulaceum serovar 1 by this analysis.     

FLYING SNAILS--HOW FAR CAN TRUNCATELLINA (PULMONATA: VERTIGINIDAE) BE BLOWN OVER THE SEA?

With populations of land snails of very small size like Vertiginidae,questions have arisen as to whether populations of relativelydistant islands in archipelagos are really isolated from eachother. Apart from other flight agencies, airborne transportof loose specimens is not improbable in stormy weather conditions.Currently, mechanisms of wind-borne transport of sand particlesover short and long distances have been intensively studied.The results are available in the literature on sediments, allowingthe calculation of probable flight distances for particles insuspension. For living snails of the Aegean species Truncatel-lina rothi,an average fall velocity of 2.6–2.7 m s^–1 has beendetermined in experiments under laboratory conditions. Applyingthese results, Truncatellina living on an island at 100 m altitudeand close to the coast could be transported up to several kilometersin heavy storms, which are not uncommon in the Aegean archipelago(Greece). This would imply that many of the Aegean islands arenot effectively isolated for minute snail species, and thatgenetic interchange between island populations is probably frequent. (Received 3 April 1996; accepted 27 January 1997)     

DNA content and p53 protein expression in ductal breast cancer

DNA content and p53 protein expression in ductal breast cancer
The DNA content of 85 ductal breast cancers of different histological grades was evaluated using static cytometry and correlated with immunocytochemical expression of p53 protein in tumour cells in cytological material. A statistically significant difference was observed between p53 protein expression and grade of malignancy ( P <0.001). The percentage of euploid tumours significantly decreased from grade I through grade II to grade III tumours ( P <0.001). Clonal DNA heterogeneity was observed in 26.6% of cases analysed and was correlated with p53 protein expression ( P <0.001). These changes probably reflect genomic alterations which may affect potential malignancy of breast cancer.