scholarly journals Pooled CRISPRi screening of the cyanobacterium Synechocystis sp. PCC 6803 for enhanced growth, tolerance, and chemical production

2019 ◽  
Author(s):  
Lun Yao ◽  
Kiyan Shabestary ◽  
Sara M. Björk ◽  
Johannes Asplund-Samuelsson ◽  
Haakan N. Joensson ◽  
...  

AbstractWe developed an inducible CRISPRi gene repression library in the cyanobacterium Synechocystis sp. PCC 6803, where all annotated genes are targeted for repression. We used the library to estimate gene fitness in multiple conditions. The library revealed several mutants with increased specific growth rates (up to 17%), and transcriptomics of these mutants revealed common upregulation of genes within photosynthetic electron flow. We challenged the library with L-lactate stress to find more tolerant mutants. Repression of the peroxiredoxin Bcp2 increased growth rate by 49% in the presence of 0.1 M L-lactate. Finally, the library was transformed into a L-lactate-secreting strain, and droplet microfluidics sorting of top producers enriched sgRNAs targeting nutrient assimilation, redox modulation, and cyclic-electron flow. Several clones showed increased productivity in batch cultivations (up to 75%). In some cases, tolerance or productivity was enhanced by partial repression of essential genes, which are difficult to access by transposon insertion.

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Lun Yao ◽  
Kiyan Shabestary ◽  
Sara M. Björk ◽  
Johannes Asplund-Samuelsson ◽  
Haakan N. Joensson ◽  
...  

AbstractCyanobacteria are model organisms for photosynthesis and are attractive for biotechnology applications. To aid investigation of genotype-phenotype relationships in cyanobacteria, we develop an inducible CRISPRi gene repression library in Synechocystis sp. PCC 6803, where we aim to target all genes for repression. We track the growth of all library members in multiple conditions and estimate gene fitness. The library reveals several clones with increased growth rates, and these have a common upregulation of genes related to cyclic electron flow. We challenge the library with 0.1 M L-lactate and find that repression of peroxiredoxin bcp2 increases growth rate by 49%. Transforming the library into an L-lactate-secreting Synechocystis strain and sorting top lactate producers enriches clones with sgRNAs targeting nutrient assimilation, central carbon metabolism, and cyclic electron flow. In many examples, productivity can be enhanced by repression of essential genes, which are difficult to access by transposon insertion.


1998 ◽  
Vol 37 (4-5) ◽  
pp. 259-262 ◽  
Author(s):  
Bjarne R. Horntvedt ◽  
Morten Rambekk ◽  
Rune Bakke

This paper presents a strategy in which mixed biological cultures are exposed to oscillating concentration levels, to improve the potential for coexistence of desired bacterial species. A mechanistic mathematical model is constructed to investigate and illustrate this strategy. This paper is focused on competition between nitrifying, denitrifying and aerobic heterotrophic bacteria in a CSTR with sludge recycle. For nitrifying and aerobic heterotrophic cultures, the effect of sinusoidal oscillations in DO levels with an amplitude of 1.0 mg/l is a 16% specific growth rate reduction compared to that at a constant DO level. The denitrifiers growth rate is increased by an average of 59%, compared to the constant DO level situation. A similar strategy has been tested in a pilot plant. It is concluded that the influence on specific growth rates is a function of the amplitude of the oscillations. The effects are greatest when concentrations fluctuate around the half saturation concentration of the rate limiting component(s).


2001 ◽  
Vol 58 (2) ◽  
pp. 386-393 ◽  
Author(s):  
John A Sweka ◽  
Kyle J Hartman

Brook trout (Salvelinus fontinalis) were held in an artificial stream to observe the influence of turbidity on mean daily consumption and specific growth rates. Treatment turbidity levels ranged from clear (<3.0 nephelometric turbidity units (NTU)) to very turbid water (> 40 NTU). Observed mean daily specific consumption rates were standardized to the mean weight of all brook trout tested. Turbidity had no significant effect on mean daily consumption, but specific growth rates decreased significantly as turbidity increased. Brook trout in turbid water became more active and switched foraging strategies from drift feeding to active searching. This switch was energetically costly and resulted in lower specific growth rates in turbid water as compared with clear water. Bioenergetics simulations were run to compare observed growth with that predicted by the model. Observed growth values fell below those predicted by the model and the difference increased as turbidity increased. Abiotic factors, such as turbidity, which bring about changes in the activity rates of fish, can have implications for the accuracy of predicted growth by bioenergetics models.


Copeia ◽  
1992 ◽  
Vol 1992 (4) ◽  
pp. 1098 ◽  
Author(s):  
Alan B. Bolten ◽  
Karen A. Bjorndal ◽  
Janice S. Grumbles ◽  
David W. Owens

2014 ◽  
Vol 17 (2) ◽  
pp. 346-363 ◽  
Author(s):  
Wout Overkamp ◽  
Onur Ercan ◽  
Martijn Herber ◽  
Antonius J. A. van Maris ◽  
Michiel Kleerebezem ◽  
...  

1995 ◽  
Vol 31 (11) ◽  
pp. 173-180 ◽  
Author(s):  
M. van der Ploeg ◽  
M. E. Dennis ◽  
M. Q. de Regt

Relative abundance of Oscillatoria cf. chalybea was monitored during May-November, 1993, in 40 ponds at four catfish farms located 50-100 km apart in west central Mississippi, USA. The occurrence of O. cf.chalybea coincided with the period that water temperatures remained above 20°C. In 70% of ponds, O. cf.chalybea was present for a period of 2-20 weeks. The alga recurred in all ponds where it had been present in 1990 and 1991. The effects of temperature and light availability on growth rate and 2-methylisoborneol (MIB) production of O. cf. chalybea were studied in continuous cultures. At 28°C, maximum specific growth rates were 0.8 d−1 (24 h light) and 0.6 d−1 (14 h light :10 h dark). Algal cells contained less MIB when adapted to the shorter light cycle than when grown under continuous light. Specific growth rate of O. cf.chalybea dropped from 0.3 to 0.1 d−1 when temperature was changed from 21 to 19.5°C (14 h light).


2021 ◽  
Author(s):  
Sevtap Tırınk ◽  
Alper Nuhoğlu ◽  
Sinan Kul

Abstract This study encompasses investigation of treatment of pistachio processing industry wastewaters in a batch reactor under aerobic conditions, calculation of kinetic parameters and comparison of different inhibition models. The mixed microorganism culture used in the study was adapted to pistachio processing industry wastewaters for nearly one month and then concentrations from 50-1000 mg L− 1 of pistachio processing industry wastewaters were added to the medium and treatment was investigated in batch experiments. The Andrews, Han-Levenspiel, Luong and Aiba biokinetic equations were chosen for the correlations between the concentration of pistachio processing industry wastewaters and specific growth rates, and the kinetic parameters in these biokinetic equations were calculated. The µmax, Ks and Ki parameters, included in the Aiba biokinetic equation providing best fit among the other equations, had values calculated as 0.25 h− 1, 19 mg L− 1, and 516 mg L− 1, respectively.


1977 ◽  
Vol 55 (10) ◽  
pp. 1756-1758 ◽  
Author(s):  
Bryan Ludwig ◽  
David A. Higgs ◽  
Ulf H. M. Fagerlund ◽  
Jack R. McBride

As part of an ongoing survey to identify hormones capable of stimulating growth in Pacific salmon, groups of underyearling coho salmon were injected with bovine (Ultralente) insulin (0.32, 1.0, 3.2, or 10 IU/kg body weight) into the peritoneal cavity either once or twice weekly for 70 days.All doses of insulin, when injected twice weekly, increased the values for specific growth rates and decreased those for food–gain ratios relative to solvent-injected controls, but the differences were not statistically significant. All doses of insulin caused a marked increase in the granulation of the pancreatic B cells. Plasma glucose concentrations in starved coho injected with 10 IU insulin/kg body weight were significantly lower than in solvent-injected controls 4 h after injection.It is concluded that proper evaluation of the effectiveness of insulin as a growth promoter for salmon requires further studies preferably using insulin preparations specific to teleosts.


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