Publicly Accessible Resources For The Cell Biology Community
To enable resource users to ascertain sequencing depth and quality underlying each variant call, we wanted to visualize the sequencing reads underlying these calls. However, there was a lack of publicly accessible visualization tools for these huge data sets. Therefore, we first designed an easily queried website front for the entire sequence variant database , allowing to quickly visualize whether a sequence of interest has either the reference sequence, unequivocally deviates from it or had issues either in the quality of the sequencing data set or in the interpretation of this data set . A description of the underlying database and the web-based visualization tool can be found in . Furthermore, from any inspected genomic region in this website, we provided a link to the sequencing read data in the publicly accessible integrative genomics viewer . Apart from allowing to visualize the basis for both calls and no calls, importantly, this integration with IGV provides for seamless visualization of the data together with the wide variety of human genome annotation tracks currently available . This enables rich data mining of 293 genome regions that are of interest to any biological study.
Figure 6: Visualization of SNPs and indels in the 293 Variant Viewer.
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Weill Cornell Medicine’s Hpsc
A team of researchers from Weill Cornell Medicine and Mount Sinai Hospital developed two novel models of humanized mice using five human embryonic stem cell lines. The first, the hPSC-LO, has human lung organoids, and the second, the hPSC-CO has human colonic organoids.
The purpose of the study, published 28 October 2020, was to study the response of human lung and colonic tissue to SARS-COV2 infection and provide in vivo platforms to facilitate drug screening of COVID-19 therapeutics.
The authors describe how they generated the hPSC-LO humanized mouse model:
Screenshot from the study:
The authors give a short description of how they generated the hPSC-CO mouse:
Humanized mice carrying hPSC-COs in vivo provide a unique platform for modeling COVID-19. In brief, hPSC-COs were transplanted under the kidney capsule of NSG mice .
Four human embyonic stem cell lines were used in the experiment. Three are identified here:
1) The RUES2 human embyronic stem cell line was derived from a frozen IVF-derived embryo . 2) The HI human embryonic cells . 3) HUES8 human embryonic stem cells
In addition, the HEK293 human embryonic kidney stem cell line was also used.
Generation And Induction Of Stable Cell Lines
For integration into the FRT docking site, cells were co-transfected with the FLP expression vector pCSFLPe and the corresponding integration vector at a ratio of 9 to 1. As the transgenes are integrated at the same chromosomal site, cell lines were prepared by pooling individual colonies. Successful integration was verified by staining the cells for -galactosidase activity. Cell lines with less than 5% blue cells were used for further experiments. For integration into the attP docking site, cells were co-transfected with the C31 integrase expression vector pchactC31hum and the corresponding integration vector at a ratio of 9 to 1. For conditional expression of transgenes, cells were cultured in 1 g/ml doxycycline or 1 M Shld1. In all experiments, cells were seeded 24 h before induction. To measure the cell proliferation rate, MTS assay was performed according to the manufacturer’s instructions .
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A Guide To Human Fetal Cell Lines From Aborted Children Used In Vaccine Development
The bodies of preborn babies have been harvested for vaccine development and testing for decades dating back as far as the 1960s. There are multiple aborted fetal cell lines currently in use, and each one can be traced back to a preborn baby.
MRC-5 is derived from the lungs of a 14-week normal male fetus. MRC-5 stands for Medical Research Council, a UK research center funded by British taxpayers, according to LifeIssues.net. The cell line was developed by J.P. Jacobs in September 1966 after an abortion was committed on a physically healthy 27-year-old woman for psychiatric reasons.
In the Journal of Biological Standardization, Jacobs wrote, The cells were derived from the lungs of a 14-week-old normal male foetus at our laboratories in 1966. The family history was genetically normal there was no indication of any congenital abnormalities and no sign of neoplastic disease at abortion.
According to Rene Leiva, M.D., author of A Brief History of Human Diploid Cell Strains, by the National Catholic Bioethics Quarterly, Jacobs also reported creating a second cell strain. MRC-9 was created from the lungs of a different aborted baby, a female approximately 15 weeks gestation in 1974. The babys 14-year-old mother allegedly had the abortion because she was unmarried.
He added, We had no donor information on 293 or what was available got lost, and this is available for PER.C6.
Hek293 Protocols: Cell Culture Transfection Protein Production
HEK 293 cells are popular for their ease of growth and transfection , making them a common cell culture in cancer research. In addition, high transfection efficiency of HEK293 cells produces exogenous proteins or viruses for pharmaceutical and biomedical research purposes. HEK-293 cells are useful for many transfection experiments, particularly the propagation of adenoviral-based and retroviral-based vectors.
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Human Embryonic Kidney 293 Cells: A Vehicle For Biopharmaceutical Manufacturing Structural Biology And Electrophysiology
State of the Art and Future Perspectives
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Famous Quotes Containing The Words List Of List Cell And/or Lines:
Do your children view themselves as successes or failures? Are they being encouraged to be inquisitive or passive? Are they afraid to challenge authority and to question assumptions? Do they feel comfortable adapting to change? Are they easily discouraged if they cannot arrive at a solution to a problem? The answers to those questions will give you a better appraisal of their education than any list of courses, grades, or test scores.
Weigh what loss your honor may sustainIf with too credent ear you list his songs,Or lose your heart, or your chaste treasure openTo his unmastered importunity.
She that but little patience knew,From childhood on, had now so muchA grey gull lost its fear and flewDown to her cell and there alit,And there endured her fingers touchAnd from her fingers ate its bit.
Was seizd by the spirit that trails in the lines underfoot,The rim, the sediment that stands for all the water and all the landof the globe.Fascinated, my eyes reverting from the south, dropt, to follow those slender windrows,Chaff, straw, splinters of wood, weeds, and the sea-gluten,Scum, scales from shining rocks, leaves of salt-lettuce, left by the tide,
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Cambridge University’s Testing Platform Of Beating Heart Muscle Cells Derived From Human Embryonic Stem Cells
This example does not involve humanized mice but is a testing model of beating cardiomycytes derived from human embryonic stem cells, called hESC-CMs. Researchers from Cambridge published a study on July 29, 2021 in which they detail how they used a testing platform of hESC-CMs to screen heart-protecting medicines for COVID-19 patients.
Human embryonic stem cell-derived cardiomyocytes were infected with SARS-CoV-2 and various drugs were tested on the cells to assess the efficacy of the inhibitors.
Screenshot from the Methods section of the Williams et al. paper:
Human embryonic stem cells from the H9 line were used. The H9 stem cell line was developed at the University of Wisconsin as described in a Science magazine article dated
6 November 1998, from thirty-six “fresh or frozen-thawed donated human embyros produced by IVF” .
H9 human embryonic stem cells were grown in colonies and differentiated into beating cardiomycytes using a process detailed in this scientific study.
Another stem cell line, the famous HEK293 human embyronic kidney stem cell line was used to develop the SARS-CoV2 pseudotyped virus, called a “pCG1-SARS-CoV-2 19 D614G HA spike plasmid.”
Screenshot from the Methods section:
Their conclusion after testing was that benztropine and DX600 are novel inhibitors of SARS-CoV-2 and could be useful in patients in whom vaccination is contraindicated.
Optimization Of The Expression System Hek293
Efficient optimization of promoter and signal peptides further enhance proteins expression. The expression can be further optimized by changing transfection protocol and cultivation time5,6.
Another approach for optimization lays in the metablism of HEK293. Understanding metabolic requirements in HEK293 significantly increases productivity through optimization of cellular metabolism Exhausted or critical substrates can be replenished and eventually toxic metabolites removed.7
This enables high throughput screening of candidate proteins to accelerate the identification of lead molecules for use in clinical trials. Thus, HEK293 cells are suitable for early stage pipeline development as well as upscaling into recombinant antibody production on industry level. HEK293 allows for greater process consistency and control of the final product quality. It can be well cultivated in serum-free media which simplifies the harvesting process.
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Human Embryonic Kidney Cells
HEK cells are the abbreviated term for Human Embryonic Kidney cells. This cell line is also referred to as HEK-293 or 293 cells. Frank Graham, who established this cell line, used to number his experiments the original HEK 293 cell clone was from his 293rd experiment, hence the full name HEK293. HEK 293 cells were generated by transfection of cultures of normal human embryonic kidney cells with sheared adenovirus 5 DNA.
HEK 293 cells have been used for many years in cell biological research as well as expression system for therapeutic proteins and viruses for gene therapy. HEK 293 immortal cell line that is comparatively easy to handle. An immortalised cell line is a population of cells from a multicellular organism which would normally not proliferate indefinitely but, due to mutation, have evaded normal cellular senescence and instead can keep undergoing division.1,2
From One To Many Hek293 And Its Derived Cell Lines
While HEK cells are cultured in many laboratories around the world, these cells are not necessarilyHEK293. Radiating outward from the original cell line, a number of derived cell lines have been established to best suit a range of applications. Introducing the viral SV40 large T antigen has createdHEK293T cells, enabling the replication of transfected plasmids with an SV40 origin of replication. This change has made HEK293T cells attractive for producing large amounts of recombinant proteins or viral particles, as high plasmid levels can be reached by transient transfections.
Both HEK293 and HEK293T represent adherent cell lines that form a stable connection with coated cell culture dishes. This limits their volume to a two-dimensional space. Therefore,HEK293F cells were selected to be cultured in suspension by adaptation, elevating them to the three-dimensional space and allowing for more efficient growth of high-density cell cultures for recombinant protein production in bioreactors. The cell lineHEK293FT further merges benefits from HEK293T and HEK293F by introducing the SV40 large T antigen into HEK293F cells as well. Next to many additional derived cell lines,HEK293SG deserves to be mentioned. Stemming from the suspension-cultured HEK293S cell line, HEK293SG are designed to produce homogeneously glycosylated biopharmaceuticals by the deletion of MGAT, a key chokepoint of the glycosylation machinery.
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Related Mammalian Cell Lines
When searching for related cell lines, ones goal is key. Different cell lines are specialized for different aspects and, for optimal results, there is no ideal cell line. For those interested in fundamental research or drug testing, other human cell lines, such as the colorectal Caco-2 cell line or the breast cancer cell line MCF-7 could be valid options. For an alternative for recombinant protein production especially for producing antibodies Chinese hamster ovary cells are among the most important cell lines. Finally, the propagation or production of viruses has to be performed in permissible cell lines for the respective virus, such as Madin-Darby Canine Kidney cells for influenza viruses.
A Summary Of Hek293 Cells And How They Are Used For Cell Biology Studies
HEK 293 cells were originally cultured by Frank Graham in 1953 and have since gone on to become one of the most widely used cell line research models after HeLa and CHO cells.
HEK 293 cells were derived from cultures of human embryonic kidney tissues and transfected with adenovirus 5 DNA. This allowed use of this viral promotor to allow an increase transcription of products such as recombinant proteins. There are now a number of HEK 293 variants which have been produced for specific applications. Some of the differences could easily be missed, so it is important to ensure the appropriate HEK 293 cell line variant is being used. A further point of interest is that HEK 293 cells cannot be used as a model to describe normal kidney cells since it was found that these cells express neuronal genes which would link them to being sourced from embryonic kidney tissue rather than adult kidney cells . Despite these caveats, these cells are still highly applicable to biotechnology applications since they are easy to work with being robust, grow quickly and easy to transfect.
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Religious Leaders And Anti
Senior Catholic leaders in the United States and Canada, along with other anti-abortion groups, have protested the use of cells derived from electively aborted human fetuses. Yesterday, Anglican Archbishop of Sydney Glenn Davies said he would likely boycott the COVID-19 vaccine being developed by Oxford University, on these ethical grounds.
The SMC asked experts to comment on the use of human embryonic cells in vaccine development.
Dr Nikki Turner, Director Immunisation Advisory Centre, University of Auckland, comments:
Human cell lines that are used for the development of some types of vaccine need to be sourced from human material. For the development of vaccines such as the rubella-containing vaccines, and chicken pox and shingles vaccines, the original source of human tissue did come from aborted fetuses.
What is very important is that the decision to terminate the fetus in these situations was unrelated to the use of the fetal material to enable sourcing of a cell line for a vaccine. Once the human cell is sourced it is then grown in the laboratories and no further sourcing is required.
No conflict of interest.
Associate Professor Helen Petousis-Harris, Vaccinologist, University of Auckland, comments:
Extended comment available on Sciblogs.
For more information about animal products and vaccines, including considerations from other religions, see the factsheet from the Immunisation Advisory Centre.
No conflict of interest.
No conflict of interest.
Why Are They Used
Before human embryonic cell lines were available, animal cell lines, such as monkey kidney, dog kidney and chicken embryo cells, were used to develop vaccines.
Between 1955 and 1963, the polio vaccine was grown in monkey kidney cells that were later found to have been infected with a virus called simian virus 40 , making vaccinated people vulnerable to SV40 infection. Modern versions of the polio vaccine are still made in a similar manner but are now extensively filtered so that the original animal cell content is removed.
The polio vaccine is also an example of a different type of vaccination from the adenovirus-based vaccines. This type of vaccine is based on an inactivated version of the poliovirus that had been grown in monkey cells. Historically, concerns about potential contamination or endemic viral content in animal cell lines encouraged the search for and use of cleaner human cell lines.
Embryonic cell lines are considered clean since they have not had time to be infected by other potentially contaminating viruses, making them safe factories for generating adenovirus-based vaccines.
Using cells from electively aborted foetuses to develop vaccines is not new. Two human embryonic cell lines called WI-38 and MRC-5, derived from electively aborted foetuses in Sweden in 1962 and the UK in 1966, respectively, have historically been used to develop weakened or inactivated virus-based vaccines against chickenpox, shingles, rubella, hepatitis A, polio and rabies.
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Cell Genomic Instability Under Selective Conditions
One of the engineering steps to derive the 293SG cell line from 293S was an EMS mutagenesis, which is introducing point mutations , followed by selection with the cytotoxic lectin ricin. From the very few resistant clones obtained, one had undetectable N-acetylglucosaminyltransferase I activity. Before the genome sequencing project, we expected to find inactivating GnTI point mutations because of the nature of the mutagenesis method that we used, but instead, a region of ~800kb at chromosome 5q35.3 has been completely deleted . This region contains the MGAT1 gene, which encodes the GnTI protein , and nine other genes unrelated to glycosylation processes. The 800-kb-deleted region is embedded in a much larger region that has undergone massive rearrangements in this clone.
Figure 5: Deletion of MGAT1 in 293SG and 293SGGD.
Selection for 293S cells without the GnTI activity of MGAT1 using EMS mutagenesis and the ricin toxin induced a 800-kb deletion at the end of chr5. This illustrates that the driving force for mutations in these cell lines are chromosomal rearrangements rather than point mutations. Simplified scheme of early N-glycan processing of glycoproteins in the Golgi apparatus. Loss of MGAT1, responsible for GnTI activity, ensures that N-glycans in the Golgi are committed to the oligomannose type. In panel a, the Y-axis represents the genomic copy number.