Chromovert® Technology is presented as a new cell engineering technology to detect and purify living cells based on gene expression.
Methods
The technology utilizes fluorogenic oligonucleotide signaling probes and flow cytometry to detect and isolate individual living cells expressing one or more transfected or endogenously-expressed genes.
Results
Results for production of cell lines expressing a diversity of ion channel and membrane proteins are presented, including heteromultimeric epithelial sodium channel (αβγ-ENaC), sodium voltage-gated ion channel 1.7 (NaV1.7-αβ1β2), four unique γ-aminobutyric acid A (GABAA) receptor ion channel subunit combinations α1β3γ2s, α2β3γ2s, α3β3γ2s and α5β3γ2s, cystic fibrosis conductance regulator (CFTR), CFTR-Δ508 and two G-protein coupled receptors (GPCRs) without reliance on leader sequences and/or chaperones. In addition, three novel plasmid-encoded sequences used to introduce 3′ untranslated RNA sequence tags in mRNA expression products and differentially-detectable fluorogenic probes directed to each are described. The tags and corresponding fluorogenic signaling probes streamline the process by enabling the multiplexed detection and isolation of cells expressing one or more genes without the need for gene-specific probes.
Conclusions
Chromovert technology is provided as a research tool for use to enrich and isolate cells engineered to express one or more desired genes.
The present work describes the reproductive biology of the white marlin (Kajikiaalbida) caught in the southwestern and equatorial Atlantic Ocean. The gonads of 924 fish were collected by observers on board Brazilian tuna longliners, between November 2004 and December 2006. The spawning season was assessed by the monthly frequency distribution of distinct stages of maturity and monthly mean female gonadosomatic index GSI. Sixty-one percent (n = 656) of the fish examined were female, with a Lower Jaw Fork Length (LJFL) between 83 and 236 cm (mean = 155.5 ± 16.63). The 268 males had a LJFL between 90 and 220 cm (mean = 152.3 ± 34.62). Although the northeastern region of Brazil does not appear to be a significant spawning area for the species, the results suggest a higher reproductive activity in the third quarter of the year. Using a Bayesian logistic model approach, length at 50% maturity was estimated at 145.04 cm (credibility interval of 95%, 143.94–146.09 cm), for females, and at 140.03 cm (credibility interval of 95%, 137.28–142.52 cm), for males. 相似文献
Splicing is catalyzed by the spliceosome, a compositionally dynamic complex assembled stepwise on pre-mRNA. We reveal links between splicing machinery components and the intrinsically disordered ciliopathy protein SANS. Pathogenic mutations in SANS/USH1G lead to Usher syndrome—the most common cause of deaf-blindness. Previously, SANS was shown to function only in the cytosol and primary cilia. Here, we have uncovered molecular links between SANS and pre-mRNA splicing catalyzed by the spliceosome in the nucleus. We show that SANS is found in Cajal bodies and nuclear speckles, where it interacts with components of spliceosomal sub-complexes such as SF3B1 and the large splicing cofactor SON but also with PRPFs and snRNAs related to the tri-snRNP complex. SANS is required for the transfer of tri-snRNPs between Cajal bodies and nuclear speckles for spliceosome assembly and may also participate in snRNP recycling back to Cajal bodies. SANS depletion alters the kinetics of spliceosome assembly, leading to accumulation of complex A. SANS deficiency and USH1G pathogenic mutations affects splicing of genes related to cell proliferation and human Usher syndrome. Thus, we provide the first evidence that splicing dysregulation may participate in the pathophysiology of Usher syndrome. 相似文献
Neurochemical Research - Gamma-decanolactone (GD) has been shown to reduce epileptic behavior in different models, inflammatory decreasing, oxidative stress, and genotoxic parameters. This study... 相似文献
Journal of Plant Growth Regulation - This study evaluated the effect of saline water irrigation (4.5 dS m−1) on growth, gas exchange and mineral nutrient content in eight melon accessions and... 相似文献
Both obesity and gestational diabetes mellitus (GDM) lead to poor maternal and fetal outcomes, including pregnancy complications, fetal growth issues, stillbirth, and developmental programming of adult-onset disease in the offspring. Increased placental oxidative/nitrative stress and reduced placental (trophoblast) mitochondrial respiration occur in association with the altered maternal metabolic milieu of obesity and GDM. The effect is particularly evident when the fetus is male, suggesting a sexually dimorphic influence on the placenta. In addition, obesity and GDM are associated with inflexibility in trophoblast, limiting the ability to switch between usage of glucose, fatty acids, and glutamine as substrates for oxidative phosphorylation, again in a sexually dimorphic manner. Here we review mechanisms underlying placental mitochondrial dysfunction: its relationship to maternal and fetal outcomes and the influence of fetal sex. Prevention of placental oxidative stress and mitochondrial dysfunction may improve pregnancy outcomes. We outline pathways to ameliorate deficient mitochondrial respiration, particularly the benefits and pitfalls of mitochondria-targeted antioxidants. 相似文献