Research on the utilization of ultra-long carbon nanotubes in lithium-ion batteries based on an environment-friendly society.

To build an environment-friendly society, clean transportation systems, and renewable energy sources play essential roles. It is critical to improve the lifetime mileage of electric vehicles' batteries for reducing the cycle life cost and carbon footprint in green transportation. In this paper, a long-life lithium-ion battery is achieved by using ultra-long carbon nanotubes (UCNTs) as a conductive agent with relatively low content (up to 0.2% wt.%) in the electrode. Ultra-long CNT could realize longer conductive path crossing active material bulks in the electrode. Meanwhile, the low content of UCNTs can help to minimize conductive agent content in electrodes and obtain higher energy density. The film resistance and electrochemical impedance spectroscopy (EIS) confirmed that the use of UCNTs could markedly enhance electronic conductivity in the battery. The battery's life and life mileage can be prolonged by almost half due to the superior electronic conductivity of UCNTs. The life cycle cost and carbon footprint are also significantly reduced, which could markedly increase economic and environmental performance.

Quantitative control of noise in mammalian gene expression by dynamic histone regulation.

Fluctuation ('noise') in gene expression is critical for mammalian cellular processes. Numerous mechanisms contribute to its origins, yet the mechanisms behind large fluctuations that are induced by single transcriptional activators remain elusive. Here, we probed putative mechanisms by studying the dynamic regulation of transcriptional activator binding, histone regulator inhibitors, chromatin accessibility, and levels of mRNAs and proteins in single cells. Using a light-induced expression system, we showed that the transcriptional activator could form an interplay with dual functional co-activator/histone acetyltransferases CBP/p300. This interplay resulted in substantial heterogeneity in H3K27ac, chromatin accessibility, and transcription. Simultaneous attenuation of CBP/p300 and HDAC4/5 reduced heterogeneity in the expression of endogenous genes, suggesting that this mechanism is universal. We further found that the noise was reduced by pulse-wide modulation of transcriptional activator binding possibly as a result of alternating the epigenetic states. Our findings suggest a mechanism for the modulation of noise in synthetic and endogenous gene expression systems.

An efficient and precise method for generating knockout cell lines based on CRISPR-Cas9 system.

Although the efficiency and versatility of CRISPR-Cas9 system has been greatly improved over conventional genome editing methods such as zinc finger or TALEN, it is still time-consuming and labor-intensive for screening knockout/knock-in cell clones due to differences of the targeted location or efficacies of guide RNAs (gRNAs). Here, we adapted a targeted knock-in strategy with CRISPR-Cas9 system and characterized the efficiency for generating single or double knockout cell lines. Specifically, a homology-arm based donor cassette consisting of genes encoding a fluorescence protein and antibiotic selection marker driven by a constitutive promoter was co-transfected with a gRNA expressing unit. Based on FACS sorting and antibiotic drug selection, positive cell clones were confirmed by genotyping and at the protein expression level. The results indicated that more than 70% of analyzed clones identified by cell sorting and selection were successfully targeted in both single and double knockout experiments. The procedure takes less than three weeks to obtain knockout cell lines. We believe that this methodology could be applicable and versatile in generating knockout cell clones with high efficiency in most cell lines.

CDR3 sequences in IgA nephropathy are shorter and exhibit reduced diversity.

Immunoglobulin (Ig) A nephropathy (IgAN) is the most common glomerulonephritis, which is characterized by the deposition of IgA antibody in the glomerulus. Systematic dissection of immune composition may contribute to a better understanding of the alternations in the immune system in IgAN. To this end, here we applied immune repertoire sequencing technology for parallel analysis of the complementary determining region 3 (CDR3) of all B cell receptors, including all five antibody subtypes (IgA, IgG, IgM, IgE and IgD), in 13 patients with IgAN and 7 healthy individuals. A significant decrease in CDR3 length was observed in the IgAN group. In particular, the JH6 family was significantly increased in IgAN. Amino acid usage was also altered in IgAN. Shannon, Simpson, Gini and Diversity 50 indices also revealed significant differences in the diversity of IgG, IgM and IgA antibodies as compared with controls. The proportions of IgA and IgG were increased, whereas IgM was decreased in IgAN. Moreover, a greater number of CDR3 sequences was shared between patients with IgAN. These findings suggest that the BCR immune repertoire is dramatically altered in IgAN, as characterized by shortened CDR3 length, as well as decreased overall diversity of CDR3.

Characterization of the cHS4 insulator in mouse embryonic stem cells.

Synthetic biology circuits are often constructed with multiple gene expression units assembled in close proximity, and they can be used to perform complex functions in embryonic stem cells (ESCs). However, mutual interference between transcriptional units has not been well studied in mouse ESCs. To assess the efficiency of insulators at suppressing promoter interference in mouse ESCs, we used an evaluation scheme in which a tunable tetracycline response element promoter is connected to a constant Nanog promoter. The chicken hypersensitive site 4 (cHS4) insulator, widely used both for enhancer blocking and for barrier insulation in vitro and in vivo, was positioned between the two expression units for assessment. By inserting the cassette into various loci of the mouse ESC genome with PiggyBac transposon, we were able to quantitatively examine the protective effect of cHS4 by gradually increasing the transcriptional activity of the tetracycline response element promoter with doxycycline and then measuring the transcriptional activity of the Nanog promoter. Our results indicate that the cHS4 insulator has minimal insulating effects on promoter interference in mouse ESCs. Further studies show that the cHS4 insulation effect may be promoter specific and related to interaction with CCCTC-binding factor-mediated loop formation. In addition, we also compared DNA transposition and transgene expression with or without the cHS4 insulator using well-established ESC reporters. The results indicate that cHS4 has no apparent effects on DNA transposition and transgene expression levels, but exerts modest protective effects on long-term transgene silencing.

PiggyBac mediated multiplex gene transfer in mouse embryonic stem cell.

PiggyBac system has been shown to have a high efficiency to mediate gene transfer. However, there are no reports on its efficiency to mediate multiplex transgenes in mouse embryonic stem cells. Here we first established an immortalized feeder cell line by introducing four antibiotic resistance genes simultaneously into the original SNL 76/7 feeder cell line utilizing the PiggyBac system. This is the feeder cell line with the most diverse types of antibiotic resistance genes reported so far, which will enable researchers to perform simultaneous multiplex gene transfer or gene targeting experiments in ES cells. With such feeder cell line, we were able to quantitatively characterize the transposition efficiency of PiggyBac system in mouse ES cells using five transposons carrying different inducible fluorescence proteins and antibiotic resistance genes, and the efficiency ranged from about 2% for one transposon to 0.5% for five transposons. The highly efficient multiplex gene transfer mediated by PiggyBac will no doubt provide researchers with more choices in biomedical research and development.

[Generation of factor VIII gene knockout mouse by tetraploid embryo complementation technology].

OBJECTIVE: Factor VIII( FVIII) gene knockout mouse model was established for further study on the treatment of hemophilia A. METHODS: Exons 16-19 of the mouse FVIII gene were knocked out by ET clone, ES homologous recombination and tetraploid embryo compensation technology. PCR, reverse transcriptase-PCR(RT-PCR) and immunohistochemistry were used to detect the transcription and translation pattern of FVIII. The phenotype of the knockout mice was analyzed by examining the activated partial thromboplastin time (APTT) and FVIII activity (FVIII:C). RESULTS: PCR, RT-PCR and immunohistochemistry confirmed that FVIII was deficient in the FVIII gene knockout mouse. The APTT results showed that FVIII-deficient mouse plasma had a prolonged clotting time compared to normal mouse plasma. The FVIII:C in heterozygous, hemizygous and homozygous mice was 80%, 8% and 10% of that in normal mice, respectively. CONCLUSION: The phenotype of the FVIII gene knockout mouse appears grossly similar to that of human with hemophilia A. Establishment of this model may promote the development of new technologies of treatment to hemophilia A.