scholarly journals Genetic engineering of sex chromosomes for batch cultivation of non-transgenic, sex-sorted males

PLoS Genetics ◽  
2020 ◽  
Vol 16 (11) ◽  
pp. e1009180
Author(s):  
Siba R. Das ◽  
Maciej Maselko ◽  
Ambuj Upadhyay ◽  
Michael J. Smanski
Keyword(s):  
Field Performance ◽  
Batch Cultivation ◽  
Genetically Engineered ◽  
Sex Selection ◽  
Batch Cultures ◽  
Genetic Method ◽  
Detection Assay ◽  
Heterogametic Sex ◽  
Genetic Constructs ◽  
Transgene Detection

The field performance of Sterile Insect Technique (SIT) is improved by sex-sorting and releasing only sterile males. This can be accomplished by resource-intensive separation of males from females by morphology. Alternatively, sex-ratio biasing genetic constructs can be used to selectively remove one sex without the need for manual or automated sorting, but the resulting genetically engineered (GE) control agents would be subject to additional governmental regulation. Here we describe and demonstrate a genetic method for the batch production of non-GE males. This method could be applied to generate the heterogametic sex (XY, or WZ) in any organism with chromosomal sex determination. We observed up to 100% sex-selection with batch cultures of more than 103 individuals. Using a stringent transgene detection assay, we demonstrate the potential of mass production of transgene free males.

scholarly journals Batch cultivation of non-transgenic males suitable for SIT programs

2019 ◽  
Author(s):  
Siba Das ◽  
Maciej Maselko ◽  
Ambuj Upadhyay ◽  
Michael J. Smanski
Keyword(s):  
Field Performance ◽  
Mass Rearing ◽  
Batch Cultivation ◽  
Genetically Engineered ◽  
Sex Selection ◽  
Batch Cultures ◽  
Detection Assay ◽  
Heterogametic Sex ◽  
Genetic Constructs ◽  
Transgene Detection

AbstractThe field performance of Sterile Insect Technique (SIT) is improved by sex-sorting and releasing only the sterile males. This can be accomplished by resource-intensive separation of males from females by morphology. Alternatively, sex-ratio biasing genetic constructs can be used to selectively kill one sex without the need for manual or automated sorting, but the resulting genetically engineered (GE) control agents would be subject to additional governmental regulation. Here we describe and demonstrate a method for the batch production of non-GE males that is applicable for sex-selective production of males suitable for genetic biocontrol programs. This method could be applied to generate the heterogametic sex (XY, or WZ) in any organism with chromosomal sex determination. We observed up to 100% sex-selection with batch cultures of more than 103 individuals. Using a stringent transgene detection assay, we demonstrate the potential of mass rearing of transgene free males.

GENETIC METHODS OF INSECT CONTROL. HISTORY AND CURRENT STATE

2021 ◽  
Vol 14 (1) ◽  
pp. 51-63
Author(s):  
A. G. Viktorov
Keyword(s):  
Invasive Species ◽  
Natural Selection ◽  
Sterile Insect Technique ◽  
Genetically Engineered ◽  
Insect Control ◽  
Genetic Method ◽  
Recessive Mutations ◽  
Current State ◽  
Invasive Mosquito ◽  
Methods Of Control

The review considers the main stages in the development of genetic method of insect control: 1) The idea of A.S. Serebrovsky, who suggested releasing males with non-viable translocations into nature; 2) "Z-lethal" method of V.A. Strunnikov for control of harmful Lepidoptera, which consists in the release of males carrying two non-allelic recessive mutations in the sex Z-chromosomes into nature; 3) Sterile insect technique (SIT) used widely in practice in the second half of the XX-th century; 4) Genetically engineered biotechnology RIDL (Release of Insects carrying a Dominant Lethal) used in practice to control the invasive mosquito Aedes aegypti , the vector of Zika viruses and yellow fever. It is generalized that the main opponent of the genetic method of insect control is natural selection, which supports females, choosing natural rather than sterile or "genetically modified" males on mating. It follows from the above that genetic methods of control can be effective only in control of invasive species have not spread widely yet.

scholarly journals Improved Production of Tryptophan in Genetically EngineeredEscherichia coliwith TktA and PpsA Overexpression

2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Tong Shen ◽  
Qing Liu ◽  
Xixian Xie ◽  
Qingyang Xu ◽  
Ning Chen
Keyword(s):  
Critical Factor ◽  
Batch Cultivation ◽  
Genetically Engineered ◽  
High Cell ◽  
Fed Batch ◽  
Acid Productivity ◽  
Genetic Modifications ◽  
Low Copy Number ◽  
Cell Densities ◽  
Fed Batch Cultivation

Intracellular precursor supply is a critical factor for amino acid productivity. In the present study,ppsAandtktAgenes were overexpressed in genetically engineeredEscherichia colito enhance the availability of two precursor substrates, phosphoenolpyruvate and erythrose-4-phosphate. The engineered strain, TRTH0709 carrying pSV709, produced 35.9 g/L tryptophan from glucose after 40 h in fed-batch cultivation. The two genes were inserted, independently or together, into a low-copy-number expression vector (pSTV28) and transferred to TRTH0709. Fed-batch fermentations at high cell densities of the recombination strains revealed that overexpression of theppsAgene alone does not significantly increase tryptophan yield. On the other hand, overexpression of thetktAgene, alone or with theppsAgene, could further improve tryptophan yield to a final tryptophan titer of 37.9 and 40.2 g/L, respectively. These results represent a 5.6% and 11.9% enhancement over the titer achieved by TRTH0709. No evident genetic modifications leading to growth impairment were observed.

scholarly journals Production of the Chiral Compound (R)-3-Hydroxybutyrate by a Genetically Engineered Methylotrophic Bacterium

2010 ◽  
Vol 76 (16) ◽  
pp. 5585-5591 ◽  
Author(s):  
Tina Hölscher ◽  
Uta Breuer ◽  
Lorenz Adrian ◽  
Hauke Harms ◽  
Thomas Maskow
Keyword(s):  
Citric Acid Cycle ◽  
Sole Source ◽  
Genetically Engineered ◽  
Methylotrophic Bacterium ◽  
Cultivation Conditions ◽  
Chiral Compound ◽  
Batch Cultures ◽  
Alternative Pathways ◽  
Intracellular Degradation ◽  
Methylobacterium Rhodesianum

ABSTRACT In this study, a methylotrophic bacterium, Methylobacterium rhodesianum MB 126, was used for the production of the chiral compound (R)-3-hydroxybutyrate (R-3HB) from methanol. R-3HB is formed during intracellular degradation of the storage polymer (R)-3-polyhydroxybutyrate (PHB). Since the monomer R-3HB does not accumulate under natural conditions, M. rhodesianum was genetically modified. The gene (hbd) encoding the R-3HB-degrading enzyme, R-3HB dehydrogenase, was inactivated in M. rhodesianum. The resulting hbd mutant still exhibited low growth rates on R-3HB as the sole source of carbon and energy, indicating the presence of alternative pathways for R-3HB utilization. Therefore, transposon mutagenesis was carried out with the hbd mutant, and a double mutant unable to grow on R-3HB was obtained. This mutant was shown to be defective in lipoic acid synthase (LipA), resulting in an incomplete citric acid cycle. Using the hbd lipA mutant, we produced 3.2 to 3.5 mM R-3HB in batch and 27 mM (2,800 mg liter−1) in fed-batch cultures. This was achieved by sequences of cultivation conditions initially favoring growth, then PHB accumulation, and finally PHB degradation.

scholarly journals Fed-Batch Cultivation and Adding Supplements to Increase Yield of β-1,3-1,4-Glucanase by Genetically Engineered Escherichia coli

Catalysts ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 269
Author(s):  
Lijuan Zhong ◽  
Zheng Liu ◽  
Yinghua Lu
Keyword(s):  
Escherichia Coli ◽  
Feeding Strategy ◽  
Batch Cultivation ◽  
Genetically Engineered ◽  
Fed Batch ◽  
Glucanase Activity ◽  
E Coli ◽  
Constant Rate ◽  
Engineered Escherichia Coli ◽  
Fed Batch Cultivation

The aim of this study was to analyze the major influence factors of culture medium on the expression level of β-1,3-1,4-glucanase, and to further develop an optimized process for the extracellular production of β-glucanase at a bioreactor scale (7 L) with a genetically engineered Escherichia coli (E. coli) JM109-pLF3. In this study, batch cultivation and fed-batch cultivation including the constant rate feeding strategy and the DO-stat (DO: Dissolved Oxygen) feeding strategy were conducted. At a 7 L bioreactor scale for batch cultivation, biomass reached 3.14 g/L and the maximum β-glucanase activity was 506.94 U/mL. Compared with batch cultivation, the addition of glycerol, complex nitrogen and complete medium during fed-batch cultivation increased the production of biomass and β-1,3-1,4-glucanase. The maximum biomass and β-glucanase activity, which were 7.67 g/L and 1680 U/mL, respectively, that is, 2.45 and 3.31 times higher than those obtained with batch cultivation, were obtained by feeding a complex nitrogen source at a constant rate of 1.11 mL/min. Therefore, these nutritional supplements and strategies can be used as a reference to enhance the production of other bioproducts from E. coli.

scholarly journals Exploiting a Y chromosome-linked Cas9 for sex selection and gene drive

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Stephanie Gamez ◽  
Duverney Chaverra-Rodriguez ◽  
Anna Buchman ◽  
Nikolay P. Kandul ◽  
Stelia C. Mendez-Sanchez ◽  
...  
Keyword(s):  
Genetic Control ◽  
Y Chromosome ◽  
Population Control ◽  
Sex Selection ◽  
Specific Gene ◽  
Gene Drive ◽  
Genetic Sexing ◽  
Animal Populations ◽  
Sexing Systems ◽  
Heterogametic Sex

AbstractCRISPR-based genetic engineering tools aimed to bias sex ratios, or drive effector genes into animal populations, often integrate the transgenes into autosomal chromosomes. However, in species with heterogametic sex chromsomes (e.g. XY, ZW), sex linkage of endonucleases could be beneficial to drive the expression in a sex-specific manner to produce genetic sexing systems, sex ratio distorters, or even sex-specific gene drives, for example. To explore this possibility, here we develop a transgenic line of Drosophila melanogaster expressing Cas9 from the Y chromosome. We functionally characterize the utility of this strain for both sex selection and gene drive finding it to be quite effective. To explore its utility for population control, we built mathematical models illustrating its dynamics as compared to other state-of-the-art systems designed for both population modification and suppression. Taken together, our results contribute to the development of current CRISPR genetic control tools and demonstrate the utility of using sex-linked Cas9 strains for genetic control of animals.

scholarly journals A Syringe-Based Biosensor to Rapidly Detect Low Levels of Escherichia Coli (ECOR13) in Drinking Water Using Engineered Bacteriophages

Sensors ◽  
2020 ◽  
Vol 20 (7) ◽  
pp. 1953 ◽  
Author(s):  
Troy C. Hinkley ◽  
Spencer Garing ◽  
Paras Jain ◽  
John Williford ◽  
Anne-Laure M. Le Ny ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Drinking Water ◽  
Membrane Filtration ◽  
Genetically Engineered ◽  
World Health ◽  
Selective Enrichment ◽  
E Coli ◽  
Detection Assay ◽  
Colony Forming Units ◽  
Health Organization

A sanitized drinking water supply is an unconditional requirement for public health and the overall prosperity of humanity. Potential microbial and chemical contaminants of drinking water have been identified by a joint effort between the World Health Organization (WHO) and the United Nations Children’s Fund (UNICEF), who together establish guidelines that define, in part, that the presence of Escherichia coli (E. coli) in drinking water is an indication of inadequate sanitation and a significant health risk. As E. coli is a nearly ubiquitous resident of mammalian gastrointestinal tracts, no detectable counts in 100 mL of drinking water is the standard used worldwide as an indicator of sanitation. The currently accepted EPA method relies on filtration, followed by growth on selective media, and requires 24–48 h from sample to results. In response, we developed a rapid bacteriophage-based detection assay with detection limit capabilities comparable to traditional methods in less than a quarter of the time. We coupled membrane filtration with selective enrichment using genetically engineered bacteriophages to identify less than 20 colony forming units (CFU) E. coli in 100 mL drinking water within 5 h. The combination of membrane filtration with phage infection produced a novel assay that demonstrated a rapid, selective, and sensitive detection of an indicator organism in large volumes of drinking water as recommended by the leading world regulatory authorities.

scholarly journals Field Performance of a Genetically Engineered Strain of Pink Bollworm

PLoS ONE ◽  
2011 ◽  
Vol 6 (9) ◽  
pp. e24110 ◽  
Author(s):  
Gregory S. Simmons ◽  
Andrew R. McKemey ◽  
Neil I. Morrison ◽  
Sinead O'Connell ◽  
Bruce E. Tabashnik ◽  
...  
Keyword(s):  
Field Performance ◽  
Genetically Engineered ◽  
Pink Bollworm ◽  
Engineered Strain

scholarly journals Increased Ethanol Productivity in Xylose-Utilizing Saccharomyces cerevisiae via a Randomly Mutagenized Xylose Reductase

2010 ◽  
Vol 76 (23) ◽  
pp. 7796-7802 ◽  
Author(s):  
David Runquist ◽  
B�rbel Hahn-H�gerdal ◽  
Maurizio Bettiga
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Parent Strain ◽  
Selection Process ◽  
Xylose Reductase ◽  
Dry Weight ◽  
Batch Cultivation ◽  
Genetically Engineered ◽  
Ethanol Productivity ◽  
Anaerobic Growth ◽  
Xylose Utilization

ABSTRACT Baker's yeast (Saccharomyces cerevisiae) has been genetically engineered to ferment the pentose sugar xylose present in lignocellulose biomass. One of the reactions controlling the rate of xylose utilization is catalyzed by xylose reductase (XR). In particular, the cofactor specificity of XR is not optimized with respect to the downstream pathway, and the reaction rate is insufficient for high xylose utilization in S. cerevisiae. The current study describes a novel approach to improve XR for ethanol production in S. cerevisiae. The cofactor binding region of XR was mutated by error-prone PCR, and the resulting library was expressed in S. cerevisiae. The S. cerevisiae library expressing the mutant XR was selected in sequential anaerobic batch cultivation. At the end of the selection process, a strain (TMB 3420) harboring the XR mutations N272D and P275Q was enriched from the library. The V max of the mutated enzyme was increased by an order of magnitude compared to that of the native enzyme, and the NADH/NADPH utilization ratio was increased significantly. The ethanol productivity from xylose in TMB 3420 was increased ∼40 times compared to that of the parent strain (0.32 g/g [dry weight {DW}] � h versus 0.007 g/g [DW] � h), and the anaerobic growth rate was increased from ∼0 h−1 to 0.08 h−1. The improved traits of TMB 3420 were readily transferred to the parent strain by reverse engineering of the mutated XR gene. Since integrative vectors were employed in the construction of the library, transfer of the improved phenotype does not require multicopy expression from episomal plasmids.

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