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Synthetic genetic devices can perform molecular computation in living bacteria, which may sense more than one environmental chemical signal, perform complex signal processing in a human-designed way, and respond in a logical manner. IMPLY is one of the four fundamental logic functions and unlike others, it is an "IF-THEN" constraint-based logic. By adopting physical hierarchy of electronics in the realm of in-cell systems chemistry, a full-spectrum transcriptional cascaded synthetic genetic IMPLY gate, which senses and integrates two environmental chemical signals, is designed, fabricated, and optimized in a single Escherichia coli cell. This IMPLY gate is successfully integrated into a 2-input-2-output integrated logic circuit and showed higher signal-decoding efficiency. Further, we showed simple application of those devices by integrating them with an inherent cellular process, where we controlled the cell morphology and color in a logical manner. To fabricate and optimize the genetic devices, a new process pipeline named NETWORK Brick is developed. This pipeline allows fast parallel kinetic optimization and reduction in the unwanted kinetic influence of one DNA module over another. A mathematical model is developed and it shows that response of the genetic devices are digital-like and are mathematically predictable. This single-cell IMPLY gate provides the fundamental constraint-based logic and completes the in-cell molecular logic processing toolbox. The work has significance in the smart biosensor, artificial in-cell molecular computation, synthetic biology, and microbiorobotics.

Biological solutions to human space travel must consider microgravity as an important component, which is unknown by the biochemical worlds on the Earth. Thus, one of the fundamental challenges of space biotechnology is to create engineered biochemical systems to integrate microgravity as a signal within molecular and cellular processes. Here we created the first molecular or biochemical microgravity sensor by creating a synthetic-small-regulatory-RNA based molecular network in E.coli, which sensed microgravity and responded by altering the expression of a target protein. We demonstrated that the design was universal, could work potentially with any promoter and against any target gene. This device was applied to target cell division process and rescue the deformed cell shape by applying microgravity. The work showed for the first time, a way to integrate microgravity as physical signals within biochemical process of a living cell in a human designed way and thus, opens a new direction in space biotechnology, space chemistry and space technology.

Bioengineering solutions to human space travel must consider microgravity as an important component. Thus, one of the fundamental challenges of space bioengineering is to create cellular microgravity responsive device, which integrate microgravity as a signal within biochemical and cellular processes. Here, we designed, fabricated and characterized the first biochemical and cellular microgravity responsive device using an engineered genetic circuit in E.coli, which responded to microgravity by changing the expression of a target enhanced green fluorescent gene (EGFP). Our device design was based on the deregulation of HfQ protein in E.coli in microgravity, which was translated through HfQ mediated silencing of EGFP by anti-EGFP synthetic small regulatory RNAs. This resulted a reduced silencing (~28 times) of the EGFP in microgravity. We demonstrated that the basic design of the device is universal in nature for E.coli, by creating multiple successful devices, where target genes (EGFP, TdTomato, and FtsZ) and the promoters (inducible and constitutive) were altered. Further, we applied this device to control the cell division process by microgravity. Here we targeted the cell division regulator FtsZ, which resulted an elongated cell shape in normal gravity and this deformed cell shape got rescued to normal one by applying microgravity. The work showed for the first time, a way to integrate microgravity as a physical signal within biochemical processes of a living cell in a human designed way and thus, may have significance in space bioengineering and synthetic biology.

Figure S1. Sequencing results for PCRed cI gene. Figure S2 Characterization of wild type (WT) cI and frame-shifted cI. Figure S3 Correlation between the dynamic range and the approximate ratio of plasmid copy number carrying frame-shifted cI and PR-EGFP construct. Figure S4 Open reading frames of wild type cI, frame-shifted cI and truncated cI started from amino acid M41. Figure S5 Generic plasmid maps constructed in this study. Figure S6 Characterization of the NOT gates repression behaviour with postulated truncated variants of cI. Table S1 Curve fitting parameter values. Table S2 Translation initiation rates for EGFP and cI calculated from RBS calculator. Table S3 List of promoters. Table S4 List of plasmids. Table S5 List of primers. (DOCX 4535 kb)

Electrical Discharge Machining (EDM) is nontraditional machining processes applied for precise machining and developing intricate geometries on work materials which are difficult to machine by conventional process. The present research work emphases on the die sinking EDM of AISI P20 tool steel, to study the effect of machining parameters such as pulse on time (POT), pulse off time (POF), discharge current (GI) and spark gap (SG) on performance response like Material removal rate (MRR), Surface Roughness (Ra) and Overcut (OC) using square-shaped Cu tool with Lateral flushing. The experimentation is performed using L27 orthogonal array and significant process parameters are ascertained using Regression analysis. The influence of the important process parameters on individual responses are detected using Cuckoo search algorithm. The present chapter is aimed at multi-response optimization i.e. higher MRR, lower Ra and minimum OC, which is conceded out using Genetic Algorithm.

Microgravity is a prominent health hazard for astronauts, yet we understand little about its effect at the molecular systems level. In this study, we have integrated a set of systems-biology tools and databases and have analysed more than 8000 molecular pathways on published global gene expression datasets of human cells in microgravity. Hundreds of new pathways have been identified with statistical confidence for each dataset and despite the difference in cell types and experiments, around 100 of the new pathways are appeared common across the datasets. They are related to reduced inflammation, autoimmunity, diabetes and asthma. We have identified downregulation of NfκB pathway via Notch1 signalling as new pathway for reduced immunity in microgravity. Induction of few cancer types including liver cancer and leukaemia and increased drug response to cancer in microgravity are also found. Increase in olfactory signal transduction is also identified. Genes, based on their expression pattern, are clustered and mathematically stable clusters are identified. The network mapping of genes within a cluster indicates the plausible functional connections in microgravity. This pipeline gives a new systems level picture of human cells under microgravity, generates testable hypothesis and may help estimating risk and developing medicine for space missions.