Transfection - Science method

Background Family with sequence similarity 134, member B (FAM134B), also known as Reticulophagy regulator 1 (RETREG1), is an ER-phagy receptor involved in ER homeostasis. Congenital mutations in the FAM134B gene have been reported to be associated with hereditary sensory and autonomic neuropathy type 2B (HSAN2B); however, the molecular differences...

TIANprep Mini Plasmid Kit is based on alkaline lysis technology followed by adsorption of DNA onto silica membrane in the presence of high salt. Plasmid DNA purified with TIANprep Mini Plasmid Kit is immediately readyfor use. Phenol extraction and ethanol precipitation are not required. High quality plasmid DNA is eluted in a small volume of Tris B...

Abstract EphrinB2 plays a critical role in tumor growth. In this study, we studied the antitumor activity of imperatorin derivative IMP-1 in renal cell carcinoma (RCC) by regulating EphrinB2 pathway.. Results showed that IMP-1 inhibited the proliferation of 786-O cells in a dose- and time-dependent manner. More importantly, knockdown and transfecti...

Background Cancer-specific adoptive T cell therapy has achieved successful milestones in multiple clinical treatments. However, the commercial production of cancer-specific T cells is often hampered by laborious cell culture procedures, the concern of retrovirus-based gene transfection, or insufficient T cell purity. Methods In this study, we deve...

Context: Impact of lost GLP-1 receptor function in human physiology. Objective: Identify coding nonsynonymous GLP1R variants in Danish individuals to link their in vitro phenotypes and clinical phenotypic associations. Methods: We sequenced GLP1R in 8,642 Danish individuals with type 2 diabetes or normal glucose tolerance and examined the abil...

Nucleic acid therapy can achieve lasting and even curative effects through gene augmentation, gene suppression, and genome editing. However, it is difficult for naked nucleic acid molecules to enter cells. As a result, the key to nucleic acid therapy is the introduction of nucleic acid molecules into cells. Cationic polymers are non-viral nucleic a...

Objective: Our previous studies showed that renal medullary sphingosine-1-phosphate receptor 1 (S1PR1) mediated sodium excretion, high salt intake increased S1PR1 level, deoxycorticosterone acetate (DOCA) blocked high salt-induced S1PR1 in the renal medulla, and that conditional knockout of S1PR1 in the collecting duct aggravated DOCA-salt hyperten...

Reverse genetic systems enable engineering of RNA virus genomes and are instrumental to study RNA virus biology. With the recent outbreak of the COVID-19 pandemic, already established methods were challenged by the large genome of SARS-CoV-2. Herein we present an elaborated strategy for the rapid and straightforward rescue of recombinant plus-stran...

Flow cytometry is a powerful quantitative assay supporting high-throughput collection of single-cell data with a high dynamic range. For flow cytometry to yield reproducible data with a quantitative relationship to the underlying biology, however, requires that 1) appropriate process controls are collected along with experimental samples, 2) these...

MicroRNA (miR/miRNA)-153, as a novel tumor-related miRNA, has been found to be aberrantly expressed in different types of cancer; however, to the best of our knowledge, the role of miR-153 in gastric cancer (GC) remains unclear. The present study demonstrated that miR-153 expression was markedly decreased in GC, including GC cell lines and culture...

Subsequently to the publication of the above article, the authors contacted the Editorial Office to explain that they had inadvertently included data from the same original source in the first row of data panels in Fig. 4B on p. 2191 (showing the results of cell migration assay experiments) to represent two differently performed experiments. Specif...

Human cytomegalovirus (HCMV) infection is associated with human glioblastoma, the most common and aggressive primary brain tumor, but the underlying infection mechanism has not been fully demonstrated. Here, we show that EphA2 was upregulated in glioblastoma and correlated with the poor prognosis of the patients. EphA2 silencing inhibits, whereas o...

Triple-negative breast cancers (TNBC) represent a pathological subtype of breast cancer, which are characterized by strong invasiveness, high metastasis rate, low survival rate, and poor prognosis, especially in patients who have developed resistance to multiline treatments. Here, we present a female patient with advanced TNBC who progressed despit...

Exosomes, a subtype of extracellular vesicles, ranging from 50 to 200 nm in diameter, and mediate cell-to-cell communication in normal biological and pathological processes. Exosomes derived from tumors have multiple functions in cancer progression, resistance, and metastasis through cancer exosome-derived tropism. However, there is no quantitative...

Transient transfection of mammalian cells using plasmid DNA is a standard method to produce adeno-associated virus (AAV) vectors allowing for flexible and scalable manufacture. Typically, three plasmids are used to encode the necessary components to facilitate vector production; however, a dual-plasmid system, termed pDG, was introduced over 2 deca...

Acute myeloid leukemia (AML) is a life-threatening aggressive malignancy of the bone marrow and has posed a great challenge to the clinic, due to a lack of fully understanding of the molecular mechanism. Histone deacetylase 1 (HDAC1) has been reported to be a therapeutic target for treating AML. Naringenin (Nar) may act as an anti-leukemic agent an...

Sanguinarine (1) is a natural product with significant pharmacological effects. However, the application of sanguinarine has been limited due to its toxic side effects and a lack of clarity regarding its molecular mechanisms. To reduce the toxic side effects of sanguinarine, its cyanide derivative (1a) was first designed and synthesized in our prev...

Infection with cagA-positive H. pylor i has been linked to an increased risk of gastric cancer. However, the precise mechanism by which cagA regulates PIM2 expression in gastric cancer is currently unknown. To address this issue, a mutant NCTC11637ΔcagA strain of H. pylori was constructed, and the effects of H. pylori/cagA on PIM2 expression in gas...

(1) Background: A premature termination codon (PTC) can be induced by a type of point mutation known as a nonsense mutation, which occurs within the coding region. Approximately 3.8% of human cancer patients have nonsense mutations of p53. However, the non-aminoglycoside drug PTC124 has shown potential to promote PTC readthrough and rescue full-len...

Osteosarcoma (OS) is a high-grade intraosseous malignancy. Twenty to thirty percent of OS patients react poorly to standard therapy with a combination of surgical resection and chemotherapy. It is necessary to find molecules that play an important role in this. This study explored the role of TRIM4 in OS chemotherapy sensitivity and malignant progr...

Background: m⁶A modification plays an important role in various biological processes and can regulate the function of non-coding RNA. Our research group found the m6A level of lncRNA TUG1 was reduced in HK-2 cells cultured with high glucose. Here we study the impact of the important component of m6A-METTL3 on TUG1 in diabetic nephropathy. Objectiv...

SARS-CoV-2 infection requires Spike protein mediating fusion between the viral and cellular membranes. The fusogenic activity of Spike requires its post-translational lipid modification by host S-acyltransferases, predominantly ZDHHC20. Previous observations indicate that SARS-CoV-2 infection augments the S-acylation of Spike when compared to trans...

The first worldwide article reporting that injections of synthetic non-replicating mRNA could be used as a vaccine, originated from a French team located in Paris was published in the European Journal of Immunology (EJI) in 1993. It relied on work conducted by several research groups in a handful of countries in the 1960s, who put forward the preci...

Cervical cancer (CC) is the most common human papillomavirus-related disease. Continuous activation of the NF-κB signaling pathway has been observed in CC. SHC binding and spindle associated 1 (SHCBP1) contributes to tumorigenesis and activation of the NF-κB pathway in multiple cancer types, while its function in CC remains unclear. In the present...

Objectives Telomerase Cajal body protein 1 (TCAB1) is closely related to the occurrence, development and prognosis of tumors, and may affect the sensitivity of tumor radiotherapy. There are no reports about the effect of TCAB1 gene expression on proliferation and apoptosis of HEPG2 cells. We plan to investigate whether silencing TCAB1 using siRNA i...

SARS-CoV-2 infection requires Spike protein mediating fusion between the viral and cellular membranes. The fusogenic activity of Spike requires its post-translational lipid modification by host S-acyltransferases, predominantly ZDHHC20. Previous observations indicate that SARS-CoV-2 infection augments the S-acylation of Spike when compared to trans...

MicroRNAs (miRNAs or miRs) play essential roles in the initiation and progression of human tumors, including cervical cancer. However, the mechanisms underlying their actions in cervical cancer remain unclear. The present study aimed to evaluate the functional role of miR‑130a‑3p in cervical cancer. Cervical cancer cells were transfected with a miR...

Transient receptor potential melastatin-like 7 (TRPM7) is a key player in various physiological and pathological processes. TRPM7 channel activity is regulated by different factors. The effects of cleavage of different domains on channel activity remain unknown. Here, we constructed several TRPM7 clones and explored the effects of truncating the mo...

In this study, we intend to explore the potential function of l-ascorbic acid in hypoxia-reoxygenation (H/R)-induced damage of CMECs and its related molecular mechanism. With different concentrations of l-ascorbic acid treatment, the proliferation, migration, inflammation and autophagy of cardiac microvascular endothelial cells (CMECs) were determi...

Introduction Our previous study has proofed the glucose sensitive gene-thioredoxin-interacting protein (TXNIP) expression was up in the placenta of the patients with gestational diabetes mellitus (GDM), but the pathological mechanisms underlying abnormal TXNIP expression in the placenta of patients with GDM is completely unclear and additional inve...

Adenosine deaminase acting on RNA ADAR1 promotes A-to-I conversion in double-stranded and structured RNAs. ADAR1 has two isoforms transcribed from different promoters: cytoplasmic ADAR1p150 is interferon-inducible while ADAR1p110 is constitutively expressed and primarily localized in the nucleus. Mutations in ADAR1 cause Aicardi - Goutières syndrom...

Gene delivery by the pulsed electric field is a promising alternative technology for nonviral transfection; however, the application of short pulses (i.e., nanosecond) is extremely limited. In this work, we aimed to show the capability to improve gene delivery using MHz frequency bursts of nanosecond pulses and characterize the potential use of gol...

RNF43 is an important negative regulator of β-catenin signaling by removing Wnt-receptors from the membrane. It is often mutated in cancers, leading to aberrant Wnt-dependent nuclear translocation of β-catenin. RNF43 has also been suggested to regulate β-catenin signaling directly within the nucleus, among other proposed nuclear functions. Given th...

Multiple sclerosis is a tissue-specific autoimmune disease of the central nervous system in which the antigen(s) remains elusive. Antibodies targeting the flotillin-1/2 complex have been described in 1-2% of the patients in a recent study. Other candidate antigens as anoctamin-2 or neurofascin-155 have been previously described in multiple sclerosi...

The long term in vitro culture of Plasmodium falciparum was successfully established by Trager and Jensen in 1976; however it largely remains unachieved for P. vivax. The major obstacle associated with Plasmodium vivax in vitro culture is its predilection for invading younger reticulocytes and the complex remodelling of invaded reticulocytes. There...

Myocardial infarction–associated transcript (MIAT) is a long noncoding RNA that plays a critical role in a variety of diseases. Accordingly, this study probed into the possible interaction mechanism between MIAT and miR-378a-5p in breast cancer. Concretely, MIAT and miR-378a-5p expressions in breast cancer tissues and cells were measured. After tra...

The human melanocortin-4 receptor plays a critical role regulating energy homeostasis. Studies on obesogenic hMC4R variants have not yet revealed how human melanocortin-4 receptor maintains body weight. Here, we identified a signaling profile for obesogenic constitutively active H76R and L250Q human melanocortin-4 receptor variants transfected in H...

Purpose: Transient receptor potential canonical 1 (TRPC1) modulates tumor growth and invasion, however, its role in tongue squamous cell carcinoma (TSCC) is unclear. The aim of this study was to explore the effect of TRPC1 knockdown on cellular function and its underlying molecular mechanism in TSCC. Methods: TSCC cell lines were transfected wit...

Introduction The purpose is to investigate the role of miR-28-5p in osteosarcoma (OS) development. Materials and Methods The expressions of miR-28-5p and URGCP in OS tissues (n = 30) and cells (MG-63 and U2OS) were discovered by q-PCR assay. MiR-28-5p mimic, sh-URGCP, pcDNA3.1-URGCP, and their controls were transfected by lipofectamine ™2000. CCK8...

The exact molecular mechanism of arsenic-induced liver injury has not been fully elucidated. The aim of the study was to investigate the potential mechanism of NaAsO2-induced cytotoxicity in BRL-3A cells and to provide a basis for the mechanism of arsenic poisoning. BRL-3A cells were treated with different doses of NaAsO2, DNMT1 inhibitor (DC_517),...

In this paper, a biologically-informed computational framework is developed to model the efficacy and to optimize the implementation of an implantable epi-retinal prosthesis that performs optogenetic stimulation through a uLED array. The developed model is capable of translating visual stimulus inputs into corresponding signals evoked in the transf...

[This corrects the article DOI: 10.3389/fgeed.2021.757540.].

Background: The E3 ubiquitin ligase casitas B-lineage lymphoma-b (CBLB) is a newly identified component of the ubiquitin-dependent protein degradation system and is thought to be an important negative regulator of immune cells. CBLB is essential for establishing a threshold for T cell activation and for regulating peripheral T cell tolerance throug...

Objective KCNQ5 encodes the voltage-gated potassium channel K V 7.5, a member of the K V 7 channel family, which conducts the M-current. This current was shown to be a potent regulator of neuronal excitability by mediating the medium and slow afterhyperpolarization. Recently, we have identified five loss-of-function variants in KCNQ5 in patients wi...

Alzheimer’s disease (AD) is one of the leading causes of death throughout the world. Z-DNA binding protein 1 (ZBP1), a DNA-related gene, is associated with inflammation, and its expression is altered in AD brain. We aimed to elucidate the exact role of ZBP1 in AD development and its potential regulatory mechanism. First, we constructed both in vivo...

E3 ubiquitin ligases play a role in protein degradation, cellular localization, and activation, and their dysregulation is associated with human diseases. Here, we present a protocol to detect IGF2BP1 ubiquitination and activation by an E3 ubiquitin ligase FBXO45. We describe steps for preparing cells and transfecting plasmids. We detail the use of...

FOXP1, known as a Forkhead-box (FOX) family protein, plays an important role in human tumorigenesis. However, the function and molecular mechanism of FOXP1 in pancreatic cancer (PC) remain unclear. Here, we report that PC patients with FOXP1 overexpression had a higher survival rate compared to patients with low- FOXP1 expression. Additionally, hig...

Primary cilia are essential sensory organelles that develop when an inhibitory cap consisting of CP110 and other proteins is eliminated. Degradation of CP110 by the ubiquitin-dependent proteasome pathway mediated by NEURL4 and HYLS1 removes the inhibitory cap. Here, we investigated the suitability of rapamycin-mediated dimerization for centriolar r...

We present here a method to culture, fix, permeabilize, and stain R1441C or VPS35 D620N MEF cells to visualize transfected Myc-RILPL1 and endogenous TMEM55b or pRab10.

To assess in vitro and in vivo tracking of iron oxide labeled stem cells transfected by lipofectamine using magnetic resonance imaging (MRI), rat dental pulp stem cells (DPSCs) were characterized, labeled with iron oxide nanoparticles, and then transfected with lipofectamine to facilitate the internalization of these nanoparticles. Cell proliferati...

Highly branched poly(β-amino ester) (HPAE) has become one of the most promising non-viral gene delivery vector candidates. When compared to other gene delivery vectors, HPAE has a broad molecular weight distribution (MWD). Despite significant efforts to optimize HPAE targeting enhanced gene delivery, the effect of different molecular weight (MW) co...

Previous studies have shown that miR-322 regulates the functions of various stem cells. However, the role and mechanism of embryonic stem cell (ESCs) differentiation into cardiomyocytes remains unknown. Celf1 plays a vital role in stem cell differentiation and may be a potential target of miR-322 in ESCs’ differentiation. We studied the function of...

The potential of therapeutically loaded nanoparticles (NPs) has been successfully demonstrated during the last decade, with NP-mediated nonviral gene delivery gathering significant attention as highlighted by the broad clinical acceptance of mRNA-based COVID-19 vaccines. A significant barrier to progress in this emerging area is the wild variabilit...

We have developed new formulations of nanohydrogels (NHGs) complexed with DNA devoid of cell toxicity, which, together with their tuned sizes, makes them of great interest for delivering DNA/RNA for foreign protein expression. Transfection results demonstrate that, unlike classical lipo/polyplexes, the new NHGs can be incubated indefinitely with ce...

Lipid-based nanoparticles have recently shown great promise, establishing themselves as the gold standard in delivering novel RNA therapeutics. However, research on the effects of storage on their efficacy, safety and stability is still lacking. Here, we explore the impact of storage temperature on two types of lipid-based nanocarriers, lipid nanop...

The goal of the study was to test the ability of a representative of the 1,4-DHP group — glutapyrone to protect DNA against damage. The compound increased the concentration of the hydroxyl radicals produced in the Fenton reaction, it did not scavenge peroxynitrite and did not protect plasmid DNA against damage by products of the Fenton reaction. On...

Polycationic carriers promise low cost and scalable gene therapy treatments, however inefficient intracellular unpacking of the genetic cargo has limited transfection efficiency. Charge-reversing polycations, which transition from cationic to neutral or negative charge, can offer targeted intracellular DNA release. We describe a new class of charge...

Despite numerous efforts over the last three decades, nucleic acid-based therapeutics still lack delivery platforms in the clinical stage. Cell-penetrating peptides (CPPs) may offer solutions as potential delivery vectors. We have previously shown that designing a "kinked" structure in the peptide backbone resulted in a CPP with efficient in vitro...

Here, we fix, permeabilize, and stain human iPSC-derived neurons for the purpose of observing and quantifying somal and axonal abundance of proteins of interest. For preceding culture of neurons, see "Protocol: Culture and transfection of iPSC-derived neurons for live-imaging of axonal cargoes."

Repairing genetic defects using exogenous DNA is a major challenge the science is currently facing. This requires the design of vectors that can effectively encapsulate, protect and target nucleic acids to specific cells safely and precisely. Here we have designed silica-based physiologically responsive particles to encapsulate, store, and transfer...

nanoparticle transfection

The development of biological methods over the past decade has stimulated great interest in the possibility to regenerate human tissues. Advances in stem cell research, gene therapy, and tissue engineering have accelerated the technology in tissue and organ regeneration. However, despite significant progress in this area, there are still several te...

Advanced intracellular delivery of proteins has profound applications in both scientific investigations and therapies. However, existing strategies relying on various chemical and physical methods, have drawbacks such as the requirement of high concentration in vitro prepared target proteins and difficulty in labeling target proteins. Developing ne...