Influences of a periodic signal on a noisy synthetic gene network

The effects of noise and a periodic signal on a synthetic gene network have been investigated. By tuning the distance of a parameter from the Hopf bifurcation point, both implicit internal signal stochastic resonance and explicit internal signal stochastic resonance can be induced by noise. Furthermore, a switch process can also be elicited. When a periodic signal is coupled to the gene network, two interesting phenomena occur with the modulation of the frequency of the signal: the effect of noise amplifying cellular signal can be inhibited or even destroyed, and "locked" coherence resonance occurs.

Microfluidic approaches for synthetic gene circuits' construction and analysis

Background:Microfluidic systems have advantages such as a high throughput,small reaction volume,and precise control of the cellular position and environment.These advantages have allowed microfluidics to be widely used in several fields of synthetic biology in recent years.Results'.In this article,we reviewed the microfluidic-based methods for synthetic biology from two aspects:the construction of synthetic gene circuits and the analysis of synthetic gene systems.We used some examples to illuminate the progresses and challenges in the steps of synthetic gene circuits construction and approaches of gene expression analysis with microfluidic systems.Conclusion:Comparing to traditional methods,microfluidic tools promise great advantages in the synthetic genetic circuit building and analysis process.Moreover,new microfluidic systems together with the mathematical modeling of synthetic circuits or consortiums are desirable to perform complex genetic circuit construction and understand the natural gene regulation in cells and population interactions better.

Cotton Plants Transformed with the Activated Chimeric Cry1Ac and API-B Genes

A chimeric gene, Bt29K, composed of coding sequences of activated Cry1Ac insecticidal protein and an endoplasm reticulum-retarding signal peptide, was synthesized. A plant expression vector containing two expression cassettes for the Bt29K and API-B genes was constructed. These two insect-resistant genes were transferred into two cotton ( Gossypium hirsutum L.) varieties ( or lines) via Agrobacterium-mediated transformation and nine homozygous transgenic cotton lines showing a mortality of 90.0% - 99.7% to cotton ballworm (Heliothis armigera) larvae and good agronomic traits were selected through six generations. Molecular biology analysis revealed that one or two copies of the insecticidal protein genes were integrated into the transgenic cotton genome and activated Cry1Ac and API-B protein expression was at a level of 0.17% and 0.09% of the total soluble protein in the transgenic cotton leaves, respectively. Comparison of the insect-resistance of the homozygous lines expressing the activated chimeric Cry1Ac and API-B with that expressing Cry1Ac only revealed that the insect-resistance of the former is apparently higher than the latter. These results also indicate that the strategy to construct a plant expression vector expressing two different insect-resistant genes reported here is reasonable.

Expression of Overlapping PCR-generated Shiga Toxin B Gene Fragment in E. Coli and Its Ascitic Polyclonal Antibody

The mature Shiga toxin B （StxB） gene was optimized and generated by overlapping PCR. Recombinant expression vector pQE40-DHFR/StxB was constructed when the gene was cloned into pQE fusion expression vector. Induced by Isopropyl β-D-thiogalactoside （IPTG）, the DHFR/StxB fusion protein was highly expressed to the level of 41.36% in E. coli MI5 cells. The 35 kDa fusion protein with a 6 His-tag was one-step purified from inclusion bodies using Ni-NTA affinity chromatography column under denaturing conditions, and was refolded by dialyzing with a decreasing urea gradient. Purified DHFR/StxB fusion protein was used to immunize Kunming mice for generating the ascitic polyclonal antibody against recombinant StxB protein by injecting sarcoma 180 cells and the titer ofascitic polyclonal antibody is up to 1： 1× 10^6 detected by the indirect enzyme linked immunosorbent assy （ELISA）. Western immunoblotting analysis revealed that the ascitic polyclonal antibody against StxB had a specific affinity for a 70 kDa shiga toxin protein of Shigella dysenteriae type 1. It is a new simple and quick method to produce a large amount of ascitic polyclonal antibody. The antibody is used to develop immunological method for detecting shiga toxin.

Engineering synthetic optogenetic networks for biomedical applications

Background： Recently, optogenetics based on genetically encoded photosensitive proteins has emerged as an innovative technology platform to revolutionize manipulation of cellular behavior through fight stimulation. It has enabled user defined control of various cellular behaviors with spatiotemporal precision and minimal invasiveness, creating unprecedented opportunities for biomedical applications. Results： This article reviews current advances in optogenetic networks designed for the treatment of human diseases. We highlight the advantages of these optogenetic networks, as well as emerging questions and future perspectives. Conclusions： Various optogenetic systems have been engineered to control biological processes at all levels using light and applied for numerous diseases, such as metabolic disorders, cancer, and immune diseases. Continued development of optogenetic modules will be necessary to precisely control of gene expression magnitude towards clinical medical practice in the context of real-world problems.

The CRISPR/Cas revolution continues: From efficient gene editing for crop breeding to plant synthetic biology

Since the discovery that nucleases of the bacterial CRISPR(clustered regularly interspaced palindromic repeat)-associated(Cas) system can be used as easily programmable tools for genome engineering,their application massively transformed different areas of plant biology. In this review, we assess the current state of their use for crop breeding to incorporate attractive new agronomical traits into specific cultivars of various crop plants. This can be achieved by the use of Cas9/12 nucleases for double-strand break induction,resulting in mutations by non-homologous recombinatr e-tion. Strategies for performing such experiments à from Rthe design of guide RNA to the use of different transformation technologies à are evaluated. Furtherweive-more, we sum up recent developments regarding the use of nuclease-deficient Cas9/12 proteins, as DNAbinding moieties for targeting different kinds of enzyme activities to specific sites within the genome. Progress in base deamination, transcriptional induction and transcriptional repression, as well as in imaging in plants, is also discussed. As different Cas9/12 enzymes are at hand, the simultaneous application of various enzyme activities, to multiple genomic sites, is now in reach to redirect plant metabolism in a multifunctional manner and pave the way for a new level of plant synthetic biology.

Genes functional identification and synthetic biology of natural products

Synthetic biology is one of the rapid developing scientific fields in recent years. Through synthetic biology,we have a better understanding of the natural synthesis process of natural products, which provides a favorable method to research the diversity of natural products, and also provides new tools for us to create new approach for producing natural products, we can synthesize compounds with different structures by artificial combination of different synthesis modules.

Chemical synthesis of a structural gene coding for trichosanthin

A structural gene (750 bp), which codes for a type I ribosome inactivating protein, trichosanthin, has been designed according to the codon usage of highly expressed gene in E. coli and chemically synthesized. In the synthesized gene, twenty-seven unique restriction sites were evenly dispersed with an average distance between two adjacent sites less than 50 bp to facilitate a systematic investigation on structure-functional relationship of this protein by site-directed mutagenesis. To synthesize it, the whole gene was divided into three large fragments (EP, PN and NH) which were assembled from several chemical synthetic oligonucleotides by enzymatic method. The assembly of both the fragment EP from six oligonucleotides (A-F) and the fragment PN from four oligomers (G-J) was catalyzed by T-4 DNA ligase in using the single stranded DNA method [Chen, H.-B. et al., Nucl. Acids Res., 18, 871(1990)]. And fragment NH was formed from three duplexes K, L and M by the classical double stranded DNA method. Finally, each fragment was cloned into vector pUC18 in succession to form the plasmid, pC0TCS, to complete the whole gene synthesis, The sequencing data for the synthetic gene coincides with the designed one.

Comparative analysis of metabolic network of pathogens

BACKGROUND： Metabolic networks are complex and system of highly connected chemical reactions and hence it needs a system level computational approach to identify the genotype- phenotype relationship. The study of essential genes and reactions and synthetic lethality of genes and reactions plays a crucial role in explaining functional links between genes and gene function predictions. METHODS： Flux balance analysis （FBA） has been developed as a powerful method for the in silico analyses of metabolic networks. In this study, we present the comparative analysis of the genomic scale metabolic networks of the four microorganisms i.e. Salmonella typhimurium, Mycobacterium tuberculosis, Staphylococcus aureus, and Helicobacter pylori. The fluxes of all reaction were obtained and the growth rate of the organism was calculated by setting the biomass reaction as the objective function. RESULTS ＆ CONCLUSIONS： The average lethality fraction of all the four organisms studied ranged from 0.2 to 0.6. It was also observed that there are very few metabolites which are highly connected. Those metabolites that are highly connected are supposed to be the ‘global players＇ similar to the hub protein in the protein - protein interaction network.