scholarly journals Improved the Agrobacterium tumefaciens-mediated transformation of cucumber by a modified the using of antibiotics and acetosyringone

2020 ◽  
Author(s):  
Li'ang Chai ◽  
Changxia Du ◽  
Huaifu Fan ◽  
Chen Liu ◽  
Yuyang Si
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Culture Medium ◽  
Early Stage ◽  
Marker Gene ◽  
Vegetable Crops ◽  
Regenerated Plants ◽  
Npt Ii ◽  
Screening Efficiency ◽  
Speed Up ◽  
Polymerase Chain

Abstract Background: Cucumber (Cucumis sativus) is one of the most important vegetable crops in the world. As conventional breeding of cucumber is very challenging, genetic engineering is an alternative option to introduce important traits such as enhanced stress resistance and nutritional value. However, the efficiency of the transformation system depends on genotypes, transformation conditions, selection agents, etc. This study aims to speed up the process of Agrobacterium-mediated transformation of cucumber. ‘ Xintai mici ’, a very popular and typical north China-type cucumber variety, was transformed with Agrobacterium GV3101. The strain carried pCAMBIA2300s plasmid, a double vector with the marker gene of neomycin phosphotransferase II ( npt II). Results: The research results indicated that cefotaxime sodium was suitable for inhibiting Agrobacterium in the stage of screening and bud elongation. Timentin was best used during rooting stage. Furthermore, 25 mg/L kanamycin was used in the early stage of screening and increased to 50 mg/L for further screening. At the bud elongation and rooting stage, 75 and 100 mg/L kanamycin was used respectively to improve the screening efficiency. In order to obtain the highest regeneration frequency of resistant buds, 50, 150, and 100 μM acetosyringone were added in the pre-culture medium, infection solution, and co-culture medium respectively. To confirm the presence of the transgenes, DNA from npt II transgenic cucumber plants was analyzed by polymerase chain reaction after transplanting resistant regenerated plants. Conclusions: We finally achieved an 8.1% conversion, which was among the highest values reported until date using cucumber ‘ Xintai mici ’. Thus an effective protocol for Agrobacterium tumefaciens -mediated genetic transformation of cucumber was optimized.

scholarly journals Improved Agrobacterium Tumefaciens-Mediated Transformation of Cucumber via Modified use of Antibiotics and Acetosyringone

2020 ◽  
Author(s):  
Li'ang Chai ◽  
Changxia Du ◽  
Huaifu Fan ◽  
Chen Liu ◽  
Yuyang Si
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Culture Medium ◽  
Early Stage ◽  
Marker Gene ◽  
Vegetable Crops ◽  
Regenerated Plants ◽  
Npt Ii ◽  
Screening Efficiency ◽  
Speed Up ◽  
Polymerase Chain

Abstract Background: Cucumber (Cucumis sativus) is one of the most important vegetable crops in the world. As conventional breeding of cucumber is very challenging, genetic engineering is an alternative option to introduce important traits such as enhanced stress resistance and nutritional value. However, the efficiency of the transformation system depends on genotypes, transformation conditions, selection agents, etc. This study aims to speed up the process of Agrobacterium-mediated transformation of cucumber. ‘Xintaimici’, a very popular and typical north China-type cucumber variety, was transformed with Agrobacterium GV3101. The strain carried the pCAMBIA2300s plasmid, a double vector with the marker gene neomycin phosphotransferase II (npt II). Results: The research results indicated that cefotaxime sodium was suitable for inhibiting Agrobacterium in the screening and bud elongation stages. Timentin was best used during the rooting stage. Furthermore, 25 mg/L kanamycin was used in the early stage of screening and increased to 50 mg/L for further screening. At the bud elongation and rooting stages, 75 and 100 mg/L kanamycin was used, respectively, to improve the screening efficiency. To obtain the highest regeneration frequency of resistant buds, 50, 150, and 100 μM acetosyringone was added in the pre-culture medium, infection solution, and co-culture medium, respectively. To confirm the presence of the transgenes, DNA from npt II transgenic cucumber plants was analysed by polymerase chain reaction after transplanting resistant regenerated plants. Conclusions: We finally achieved an 8.1% conversion, which is among the highest values reported to date using the cucumber ‘Xintaimici’. Thus, an effective protocol for Agrobacterium tumefaciens-mediated genetic transformation of cucumber was optimized.

scholarly journals Improved Agrobacterium Tumefaciens-Mediated Transformation of Cucumber via Modified use of Antibiotics and Acetosyringone

2020 ◽  
Author(s):  
Li'ang Chai ◽  
Changxia Du ◽  
Huaifu Fan ◽  
Chen Liu ◽  
Yuyang Si
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Culture Medium ◽  
Early Stage ◽  
Marker Gene ◽  
Vegetable Crops ◽  
Regenerated Plants ◽  
Npt Ii ◽  
Screening Efficiency ◽  
Speed Up ◽  
Polymerase Chain

Abstract Background: Cucumber (Cucumis sativus) is one of the most important vegetable crops in the world. As conventional breeding of cucumber is very challenging, genetic engineering is an alternative option to introduce important traits such as enhanced stress resistance and nutritional value. However, the efficiency of the transformation system depends on genotypes, transformation conditions, selection agents, etc. This study aims to speed up the process of Agrobacterium-mediated transformation of cucumber. ‘Xintaimici’, a very popular and typical north China-type cucumber variety, was transformed with Agrobacterium GV3101. The strain carried the pCAMBIA2300s plasmid, a double vector with the marker gene neomycin phosphotransferase II (npt II). Results: The research results indicated that cefotaxime sodium was suitable for inhibiting Agrobacterium in the screening and bud elongation stages. Timentin was best used during the rooting stage. Furthermore, 25 mg/L kanamycin was used in the early stage of screening and increased to 50 mg/L for further screening. At the bud elongation and rooting stages, 75 and 100 mg/L kanamycin was used, respectively, to improve the screening efficiency. To obtain the highest regeneration frequency of resistant buds, 50, 150, and 100 μM acetosyringone was added in the pre-culture medium, infection solution, and co-culture medium, respectively. To confirm the presence of the transgenes, DNA from npt II transgenic cucumber plants was analysed by polymerase chain reaction after transplanting resistant regenerated plants. Conclusions: We finally achieved an 8.1% conversion, which is among the highest values reported to date using the cucumber ‘Xintaimici’. Thus, an effective protocol for Agrobacterium tumefaciens-mediated genetic transformation of cucumber was optimized.

scholarly journals High-throughput microbioreactor provides a capable tool for early stage bioprocess development

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Mathias Fink ◽  
Monika Cserjan-Puschmann ◽  
Daniela Reinisch ◽  
Gerald Striedner
Keyword(s):  
High Throughput ◽  
Process Development ◽  
Early Stage ◽  
Explanatory Power ◽  
Batch Mode ◽  
E Coli ◽  
Speed Up ◽  
Bioreactor System ◽  
Cell Densities ◽  
Microbial Systems

AbstractTremendous advancements in cell and protein engineering methodologies and bioinformatics have led to a vast increase in bacterial production clones and recombinant protein variants to be screened and evaluated. Consequently, an urgent need exists for efficient high-throughput (HTP) screening approaches to improve the efficiency in early process development as a basis to speed-up all subsequent steps in the course of process design and engineering. In this study, we selected the BioLector micro-bioreactor (µ-bioreactor) system as an HTP cultivation platform to screen E. coli expression clones producing representative protein candidates for biopharmaceutical applications. We evaluated the extent to which generated clones and condition screening results were transferable and comparable to results from fully controlled bioreactor systems operated in fed-batch mode at moderate or high cell densities. Direct comparison of 22 different production clones showed great transferability. We observed the same growth and expression characteristics, and identical clone rankings except one host-Fab-leader combination. This outcome demonstrates the explanatory power of HTP µ-bioreactor data and the suitability of this platform as a screening tool in upstream development of microbial systems. Fast, reliable, and transferable screening data significantly reduce experiments in fully controlled bioreactor systems and accelerate process development at lower cost.

scholarly journals Early Confirmation of Mycoplasma pneumoniae Infection by Two Short-Term Serologic IgM Examination

Diagnostics ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 353
Author(s):  
Ha Eun Jeon ◽  
Hyun Mi Kang ◽  
Eun Ae Yang ◽  
Hye Young Han ◽  
Seung Beom Han ◽  
...  
Keyword(s):  
Mycoplasma Pneumoniae ◽  
Early Stage ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Short Term ◽  
Serologic Tests ◽  
Polymerase Chain ◽  
Positive Correlations ◽  
Patient Selection Bias ◽  
Mycoplasma Pneumoniae Infection

The aim of the present study is to re-evaluate the clinical application of two-times serologic immunoglobulin M (IgM) tests using microparticle agglutination assay (MAA), an enzyme-linked immunosorbent assay (ELISA), and polymerase chain reaction (PCR) assay in diagnosing Mycoplasma pneumoniae (MP) infection. A retrospective analysis of 62 children with MP pneumonia during a recent epidemic (2019–2020) was conducted. The MAA and ELISA immunoglobulin M (IgM) and IgG measurements were conducted twice at admission and around discharge, and MP PCR once at presentation. Diagnostic rates in each test were calculated at presentation and at discharge. The seroconverters were 39% (24/62) of patients tested by MAA and 29% (18/62) by ELISA. At presentation, the diagnostic positive rates of MAA, ELISA, and PCR tests were 61%, 71%, and 52%, respectively. After the second examination, the rates were 100% in both serologic tests. There were positive correlations between the titers of MAA and the IgM values of ELISA. The single serologic IgM or PCR tests had limitations to select patients infected with MP in the early stage. The short-term, paired IgM serologic tests during hospitalization can reduce patient-selection bias in MP infection studies.

scholarly journals Cytokinin Mediated Increased In Vitro Production of Secondary Metabolites with Special Reference to Solasodine in Solanum erianthum

2021 ◽  
Vol 8 (02) ◽  
pp. e62-e68
Author(s):  
Jeeta Sarkar ◽  
Nirmalya Banerjee
Keyword(s):  
Secondary Metabolites ◽  
High Performance ◽  
Culture Medium ◽  
In Vitro Production ◽  
Leaf Extracts ◽  
Plant Parts ◽  
Regenerated Plants ◽  
Vitro Production ◽  
Regenerated Plantlets

AbstractSteroid alkaloid solasodine is a nitrogen analogue of diosgenin and has great importance in the production of steroidal medicines. Solanum erianthum D. Don (Solanaceae) is a good source of solasodine. The aim of this study was to evaluate the effect of different cytokinins on the production of secondary metabolites, especially solasodine in the in vitro culture of S. erianthum. For solasodine estimation, field-grown plant parts and in vitro tissues were extracted thrice and subjected to high-performance liquid Chromatography. Quantitative analysis of different secondary metabolites showed that the amount was higher in the in vitro regenerated plantlets compared to callus and field-grown plants. The present study critically evaluates the effect of the type of cytokinin used in the culture medium on solasodine accumulation in regenerated plants. The highest solasodine content (46.78±3.23 mg g-1) was recorded in leaf extracts of the in vitro grown plantlets in the presence of 6-γ,γ-dimethylallylamino purine in the culture medium and the content was 3.8-fold higher compared to the mother plant.

Acrolein produced from polyamines as one of the uraemic toxins

2003 ◽  
Vol 31 (2) ◽  
pp. 371-374 ◽  
Author(s):  
K. Sakata ◽  
K. Kashiwagi ◽  
S. Sharmin ◽  
S. Ueda ◽  
K. Igarashi
Keyword(s):  
Renal Failure ◽  
Chronic Renal Failure ◽  
Oxidase Activity ◽  
Culture Medium ◽  
Cellular Proliferation ◽  
Amine Oxidase ◽  
Early Stage ◽  
Chronic Glomerulonephritis ◽  
Toxic Compound ◽  
Amine Oxidase Activity

It is well known that the addition of spermine or spermidine to culture medium containing ruminant serum inhibits cellular proliferation. This effect is caused by the products of oxidation of polyamines that are generated by serum amine oxidase. Among the products, we found that acrolein is a major toxic compound produced from spermine and spermidine by amine oxidase. We then analysed the level of polyamines (putrescine, spermidine and spermine) and amine oxidase activity in plasma of patients with chronic renal failure. It was found that the levels of putrescine and the amine oxidase activity were increased, whereas spermidine and spermine were decreased in plasma of patients with chronic renal failure. The levels of free and protein-conjugated acrolein were also increased in plasma of patients with chronic renal failure. An increase in putrescine, amine oxidase and acrolein in plasma was observed in all cases such as diabetic nephropathy, chronic glomerulonephritis and nephrosclerosis. These results suggest that acrolein is produced during the early stage of nephritis through kidney damage and also during uraemia through accumulation of polyamines in blood due to the decrease in their excretion into urine.

scholarly journals AI-Empowered Computational Examination of Chest Imaging for COVID-19 Treatment: A Review

2021 ◽  
Vol 4 ◽  
Author(s):  
Hanqiu Deng ◽  
Xingyu Li
Keyword(s):  
False Negative ◽  
Screening Tools ◽  
Polymerase Chain Reaction Test ◽  
Test Results ◽  
First Case ◽  
Lung Scan ◽  
Screening Efficiency ◽  
Polymerase Chain ◽  
Chain Reaction Test ◽  
Full Swing

Since the first case of coronavirus disease 2019 (COVID-19) was discovered in December 2019, COVID-19 swiftly spread over the world. By the end of March 2021, more than 136 million patients have been infected. Since the second and third waves of the COVID-19 outbreak are in full swing, investigating effective and timely solutions for patients’ check-ups and treatment is important. Although the SARS-CoV-2 virus-specific reverse transcription polymerase chain reaction test is recommended for the diagnosis of COVID-19, the test results are prone to be false negative in the early course of COVID-19 infection. To enhance the screening efficiency and accessibility, chest images captured via X-ray or computed tomography (CT) provide valuable information when evaluating patients with suspected COVID-19 infection. With advanced artificial intelligence (AI) techniques, AI-driven models training with lung scans emerge as quick diagnostic and screening tools for detecting COVID-19 infection in patients. In this article, we provide a comprehensive review of state-of-the-art AI-empowered methods for computational examination of COVID-19 patients with lung scans. In this regard, we searched for papers and preprints on bioRxiv, medRxiv, and arXiv published for the period from January 1, 2020, to March 31, 2021, using the keywords of COVID, lung scans, and AI. After the quality screening, 96 studies are included in this review. The reviewed studies were grouped into three categories based on their target application scenarios: automatic detection of coronavirus disease, infection segmentation, and severity assessment and prognosis prediction. The latest AI solutions to process and analyze chest images for COVID-19 treatment and their advantages and limitations are presented. In addition to reviewing the rapidly developing techniques, we also summarize publicly accessible lung scan image sets. The article ends with discussions of the challenges in current research and potential directions in designing effective computational solutions to fight against the COVID-19 pandemic in the future.

scholarly journals Double Haploid Development and Assessment of Androgenic Competence of Balkan Pepper Core Collection in Bulgaria

Plants ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 2414
Author(s):  
Stanislava Grozeva ◽  
Gancho Pasev ◽  
Vesela Radeva-Ivanova ◽  
Velichka Todorova ◽  
Valentina Ivanova ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Double Haploid ◽  
Genetic Improvement ◽  
Core Collection ◽  
Culture Medium ◽  
Doubled Haploids ◽  
Caps Marker ◽  
Regenerated Plants ◽  
Direct Embryogenesis ◽  
Pepper Breeding

This study was designed to assess the androgenic potential of 180 pepper accessions and 11 progenies (four F1 and seven BC) possessing PMMoV resistance in order to complement an ongoing pepper breeding program. The experiment was carried out in 10 replications with 20 anthers for each accession in two different induction mediums from 2017 to 2019. The highest androgenic response was observed in culture medium 17-2 but differences between two mediums were nonsignificant. From a total of 191 genotypes, 102 genotypes expressed a potential for direct embryogenesis. Embryo induction was seen to be genotype-dependent and decreased in the following order: Pumpkin > Conical > Bell or blocky > Round > Elongate as the most responsive genotypes with over 10% reacted anthers being observed in CAPS-23, CAPS-29, CAPS-127, CAPS-157, CAPS-169, F1 and BC 887 derived from CAPS-23. The number of regenerated plants was higher in the conical group and least in the round varietal group. Regenerated plants were examined visually and by flow cytometry for identification of spontaneous doubled haploids (DH) and haploids. Those originating from F1 and BC progenies were additionally evaluated by a CAPS marker targeting L4 allele for resistance against PMMoV. Obtained results revealed two groups consisting of homozygous susceptible and resistant plants. Therefore, use of anther culture in ongoing breeding will greatly facilitate the pepper genetic improvement.

scholarly journals Optimized switch-level soft error detection based on advanced switch-level models

2021 ◽  
Author(s):  
Jalal Mohammad Chikhe
Keyword(s):  
Error Detection ◽  
Early Stage ◽  
Soft Errors ◽  
Logic Gate ◽  
Soft Error ◽  
Combinational Circuits ◽  
Single Event Transient ◽  
New Techniques ◽  
Speed Up ◽  
Logic Threshold

Due to the reduction of transistor size, modern circuits are becoming more sensitive to soft errors. The development of new techniques and algorithms targeting soft error detection are important as they allow designers to evaluate the weaknesses of the circuits at an early stage of the design. The project presents an optimized implementation of soft error detection simulator targeting combinational circuits. The developed simulator uses advanced switch level models allowing the injection of soft errors caused by single event-transient pulses with magnitudes lesser than the logic threshold. The ISCAS'85 benchmark circuits are used for the simulations. The transients can be injected at drain, gate, or inputs of logic gate. This gives clear indication of the importance of transient injection location on the fault coverage. Furthermore, an algorithm is designed and implemented in this work to increase the performance of the simulator. This optimized version of the simulator achieved an average speed-up of 310 compared to the non-algorithm based version of the simulator.

scholarly journals Detection of Oil Palm Root Penetration by Agrobacterium-Mediated Transformed Ganoderma boninense, Expressing Green Fluorescent Protein

2017 ◽  
Vol 107 (4) ◽  
pp. 483-490 ◽  
Author(s):  
Nisha Govender ◽  
Mui-Yun Wong
Keyword(s):  
Oil Palm ◽  
Fluorescent Protein ◽  
Fusion Gene ◽  
Early Stage ◽  
Marker Gene ◽  
Induction Medium ◽  
Root Penetration ◽  
Ganoderma Boninense ◽  
Agrobacterium Strain ◽  
Green Fluorescent

A highly efficient and reproducible Agrobacterium-mediated transformation protocol for Ganoderma boninense was developed to facilitate observation of the early stage infection of basal stem rot (BSR). The method was proven amenable to different explants (basidiospore, protoplast, and mycelium) of G. boninense. The transformation efficiency was highest (62%) under a treatment combination of protoplast explant and Agrobacterium strain LBA4404, with successful expression of an hyg marker gene and gus-gfp fusion gene under the control of heterologous p416 glyceraldehyde 3-phosphate dehydrogenase promoter. Optimal transformation conditions included a 1:100 Agrobacterium/explant ratio, induction of Agrobacterium virulence genes in the presence of 250 μm acetosyringone, co-cultivation at 22°C for 2 days on nitrocellulose membrane overlaid on an induction medium, and regeneration of transformants on potato glucose agar prepared with 0.6 M sucrose and 20 mM phosphate buffer. Evaluated transformants were able to infect root tissues of oil palm plantlets with needle-like microhyphae during the penetration event. The availability of this model pathogen system for BSR may lead to a better understanding of the pathogenicity factors associated with G. boninense penetration into oil palm roots.

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