Yanyi Huang

Highly sensitive fiber Bragg grating refractive index sensors

We combine fiber Bragg grating (FBG) technology with a wet chemical etch-erosion procedure and demonstrate two types of refractive index sensors using single-mode optical fibers. The first index sensor device is an etch-eroded single FBG with a radius of 3 μm, which is used to measure the indices of four different liquids. The second index sensor device is an etch-eroded fiber Fabry-Perot interferometer (FFPI) with a radius of ~1.5 μm and is used to measure the refractive indices of isopropyl alcohol solutions of different concentrations. Due to its narrower resonance spectral feature, the FFPI sensor has a higher sensitivity than the FBG sensor and can detect an index variation of 1.4 X 10(-5). Since we can measure the reflection signal, these two types of sensors can be fabricated at the end of a fiber and used as point sensors.

High-throughput screening of a CRISPR/Cas9 library for functional genomics in human cells

Targeted genome editing technologies are powerful tools for studying biology and disease, and have a broad range of research applications. In contrast to the rapid development of toolkits to manipulate individual genes, large-scale screening methods based on the complete loss of gene expression are only now beginning to be developed. Here we report the development of a focused CRISPR/Cas-based (clustered regularly interspaced short palindromic repeats/CRISPR-associated) lentiviral library in human cells and a method of gene identification based on functional screening and high-throughput sequencing analysis. Using knockout library screens, we successfully identified the host genes essential for the intoxication of cells by anthrax and diphtheria toxins, which were confirmed by functional validation. The broad application of this powerful genetic screening strategy will not only facilitate the rapid identification of genes important for bacterial toxicity but will also enable the discovery of genes that participate in other biological processes.

Fabrication of functional microstructured optical fibers through a selective-filling technique

We develop a method for the fabrication of functional microstructured optical fibers (MOFs) by selectively filling the air holes with liquid phase materials, where we utilize the dependence of filling speed on the size of the air holes. As a demonstration, we construct a hybrid MOF by filling the center hollow core of a triangular lattice photonic crystal fiber with dye-doped curable polymer, and experimentally observe the two-photon fluorescence from the hybrid MOF.

Microfluidic single-cell whole-transcriptome sequencing.

Significance RNA sequencing of single cells enables measurement of biological variation in heterogeneous cellular populations and dissection of transcriptome complexity that is masked in ensemble measurements of gene expression. The low quantity of RNA in a single cell, however, hinders efficient and consistent reverse transcription and amplification of cDNA, limiting accuracy and obscuring biological variation with high technical noise. We developed a microfluidic approach to prepare cDNA from single cells for high-throughput transcriptome sequencing. The microfluidic platform facilitates single-cell manipulation, minimizes contamination, and furthermore, provides improved detection sensitivity and measurement precision, which is necessary for differentiating biological variability from technical noise. Single-cell whole-transcriptome analysis is a powerful tool for quantifying gene expression heterogeneity in populations of cells. Many techniques have, thus, been recently developed to perform transcriptome sequencing (RNA-Seq) on individual cells. To probe subtle biological variation between samples with limiting amounts of RNA, more precise and sensitive methods are still required. We adapted a previously developed strategy for single-cell RNA-Seq that has shown promise for superior sensitivity and implemented the chemistry in a microfluidic platform for single-cell whole-transcriptome analysis. In this approach, single cells are captured and lysed in a microfluidic device, where mRNAs with poly(A) tails are reverse-transcribed into cDNA. Double-stranded cDNA is then collected and sequenced using a next generation sequencing platform. We prepared 94 libraries consisting of single mouse embryonic cells and technical replicates of extracted RNA and thoroughly characterized the performance of this technology. Microfluidic implementation increased mRNA detection sensitivity as well as improved measurement precision compared with tube-based protocols. With 0.2 M reads per cell, we were able to reconstruct a majority of the bulk transcriptome with 10 single cells. We also quantified variation between and within different types of mouse embryonic cells and found that enhanced measurement precision, detection sensitivity, and experimental throughput aided the distinction between biological variability and technical noise. With this work, we validated the advantages of an early approach to single-cell RNA-Seq and showed that the benefits of combining microfluidic technology with high-throughput sequencing will be valuable for large-scale efforts in single-cell transcriptome analysis.

Genome-wide functional screening of miR-23b as a pleiotropic modulator suppressing cancer metastasis

miRNA globally deregulates human carcinoma. A critical open question is how many miRNAs functionally participate in cancer development, particularly in metastasis. We systematically evaluate the capability of all known human miRNAs to regulate certain metastasis-relevant cell behaviours. To perform the high-throughput screen of miRNAs, which regulate cell migration, we developed a novel self-assembled cell microarray. Here we show that over 20% of miRNAs have migratory regulation activity in diverse cell types, indicating a general involvement of miRNAs in migratory regulation. MiR-23b, which is downregulated in human colon cancer samples, potently mediates the multiple steps of metastasis, including tumour growth, invasion and angiogenesis in vivo. It regulates a cohort of prometastatic targets, including FZD7 or MAP3k1. These findings provide new insight into the physiological and potential therapeutic importance of miRNAs as a new class of functional modulators.

Single-cell triple omics sequencing reveals genetic, epigenetic, and transcriptomic heterogeneity in hepatocellular carcinomas

Single-cell genome, DNA methylome, and transcriptome sequencing methods have been separately developed. However, to accurately analyze the mechanism by which transcriptome, genome and DNA methylome regulate each other, these omic methods need to be performed in the same single cell. Here we demonstrate a single-cell triple omics sequencing technique, scTrio-seq, that can be used to simultaneously analyze the genomic copy-number variations (CNVs), DNA methylome, and transcriptome of an individual mammalian cell. We show that large-scale CNVs cause proportional changes in RNA expression of genes within the gained or lost genomic regions, whereas these CNVs generally do not affect DNA methylation in these regions. Furthermore, we applied scTrio-seq to 25 single cancer cells derived from a human hepatocellular carcinoma tissue sample. We identified two subpopulations within these cells based on CNVs, DNA methylome, or transcriptome of individual cells. Our work offers a new avenue of dissecting the complex contribution of genomic and epigenomic heterogeneities to the transcriptomic heterogeneity within a population of cells.

Piezoelectric Potential Gated Field-Effect Transistor Based on a Free-Standing ZnO Wire

We report an external force triggered field-effect transistor based on a free-standing piezoelectric fine wire (PFW). The device consists of an Ag source electrode and an Au drain electrode at two ends of a ZnO PFW, which were separated by an insulating polydimethylsiloxane (PDMS) thin layer. The working principle of the sensor is proposed based on the piezoelectric potential gating effect. Once subjected to a mechanical impact, the bent ZnO PFW cantilever creates a piezoelectric potential distribution across it width at its root and simultaneously produces a local reverse depletion layer with much higher donor concentration than normal, which can dramatically change the current flowing from the source electrode to drain electrode when the device is under a fixed voltage bias. Due to the free-standing structure of the sensor device, it has a prompt response time less than 20 ms and quite high and stable sensitivity of 2%/microN. The effect from contact resistance has been ruled out.

Ultra-large bandwidth hollow-core guiding in all-silica bragg fibers with nano-supports

We demonstrate a new class of hollow-core Bragg fibers that are composed of concentric cylindrical silica rings separated by nanoscale support bridges. We theoretically predict and experimentally observe hollow-core confinement over an octave frequency range. The bandwidth of bandgap guiding in this new class of Bragg fibers exceeds that of other hollow-core fibers reported in the literature. With only three rings of silica cladding layers, these Bragg fibers achieve propagation loss of the order of 1 dB/m.

Measuring rapid enzymatic kinetics by electrochemical method in droplet-based microfluidic devices with pneumatic valves.

This paper describes a droplet-based microfluidic chip with pneumatic valves for measuring millisecond enzyme kinetics using amperometric detection method. Aqueous streams containing reactants were injected to an oil flow to form droplets, and each droplet represented one microreactor. Pneumatic valves were used to control the moving distance and in turn the reaction time of the droplets. The reaction time was also fine-tuned by varying the flow rate of the droplets in microchannels. A complete Michaelis-Menten kinetics of catalase was successfully measured by amperometric method in a single-run experiment, and the total consumption of reagents was less than 50 microL. In the current experiment, the best time resolution was about 0.05 s, and the reaction time measured was from 0.05 to 25 s. This microfluidic system is applicable to many biochemical reactions, as long as one of the reactants or products is electrochemically active. With appropriate quenching method at the outlet, various detection methods can be integrated into the microfluidic system, further extending the application of the combination of pneumatic valves and droplets in microchannels.

Single-cell RNA-seq transcriptome analysis of linear and circular RNAs in mouse preimplantation embryos

Circular RNAs (circRNAs) are a new class of non-polyadenylated non-coding RNAs that may play important roles in many biological processes. Here we develop a single-cell universal poly(A)-independent RNA sequencing (SUPeR-seq) method to sequence both polyadenylated and non-polyadenylated RNAs from individual cells. This method exhibits robust sensitivity, precision and accuracy. We discover 2891 circRNAs and 913 novel linear transcripts in mouse preimplantation embryos and further analyze the abundance of circRNAs along development, the function of enriched genes, and sequence features of circRNAs. Our work is key to deciphering regulation mechanisms of circRNAs during mammalian early embryonic development.