A5114377132- Fungal and yeast genetics research
- CRISPR and Genetic Engineering
- RNA and protein synthesis mechanisms
- DNA Repair Mechanisms
- Chromosomal and Genetic Variations
- Genomics and Chromatin Dynamics
- Microbial Metabolic Engineering and Bioproduction
- Genomics and Phylogenetic Studies
- DNA and Biological Computing
- Biotechnology and Related Fields
- Biomedical and Engineering Education
- Bacterial Genetics and Biotechnology
- Genetics, Bioinformatics, and Biomedical Research
- Plant biochemistry and biosynthesis
- DNA and Nucleic Acid Chemistry
- Advanced Proteomics Techniques and Applications
- Receptor Mechanisms and Signaling
- Advanced biosensing and bioanalysis techniques
- Microtubule and mitosis dynamics
- Science, Research, and Medicine
- Viral Infectious Diseases and Gene Expression in Insects
- Ubiquitin and proteasome pathways
- Gene expression and cancer classification
- Genomic variations and chromosomal abnormalities
- Genetics and Neurodevelopmental Disorders
Institute for Systems Biology
2017-2023
Johns Hopkins University
2014-2017
NYU Langone Health
2014-2017
Johns Hopkins Medicine
2017
High Throughput Biology (United States)
2014-2015
New York University
2015
Tel Aviv University
2009-2013
Designer Chromosome One of the ultimate aims synthetic biology is to build designer organisms from ground up. Rapid advances in DNA synthesis has allowed assembly complete bacterial genomes. Eukaryotic organisms, with their generally much larger and more complex genomes, present an additional challenge biologists. Annaluru et al. (p. 55 , published online 27 March) designed a eukaryotic chromosome based on yeast III. The chromosome, shorn destabilizing transfer RNA genes transposons, ∼14%...
The Sc2.0 project is building a eukaryotic synthetic genome from scratch. A major milestone has been achieved with all individual chromosomes assembled. Here, we describe the consolidation of multiple using advanced endoreduplication intercrossing tRNA expression cassettes to generate strain 6.5 chromosomes. 3D chromosome organization and transcript isoform profiles were evaluated Hi-C long-read direct RNA sequencing. We developed CRISPR Directed Biallelic URA3-assisted Genome Scan, or...
Extrachomosomal circular DNA (eccDNA) is ubiquitous in eukaryotic organisms and was detected every organism tested, including humans. A two-dimensional gel electrophoresis facilitates the detection of eccDNA preparations genomic DNA. Using this technique we have previously demonstrated that most consists exact multiples chromosomal tandemly repeated DNA, both coding genes satellite Here report occurrence tested human cell line. It has heterogeneous mass ranging from less than 2 kb to over 20...
We have developed a method for assembling genetic pathways expression in Saccharomyces cerevisiae. Our pathway assembly method, called VEGAS (Versatile system), exploits the native capacity of S. cerevisiae to perform homologous recombination and efficiently join sequences with terminal homology. In workflow, homology between adjacent genes vector is encoded by ‘VEGAS adapter’ (VA) sequences, which are orthogonal sequence respect yeast genome. Prior cerevisiae, each gene assigned an...
We have adapted the Golden Gate DNA assembly method to of transcription units (TUs) for yeast Saccharomyces cerevisiae, in a we call (yGG). yGG allows easy TUs consisting promoters (PRO), coding sequences (CDS), and terminators (TER). Carefully designed overhangs exposed by digestion with type IIS restriction enzyme enable virtually seamless that, principle, contain all information necessary express gene interest yeast. also describe versatile set acceptor vectors be used TU assembly. These...
Significance The advance of biotechnology opens up greater possibilities bioterror and bioerror. Here, we propose multiplexed safeguard switches rooted in the development foundational genomic, regulatory, metabolic technologies. Safeguard can be regulated by submicromolar small molecule(s) combined a modular fashion. resulting strains show high fitness low reversion rates. Moreover, two distinct classes are orthogonal, providing potential fail-safe mechanism. technologies provide practical...
Engineering multicellularity is one of the next breakthroughs for Synthetic Biology. A key bottleneck to building multicellular systems lack a scalable signaling language with large number interfaces that can be used simultaneously. Here, we present modular, scalable, intercellular in yeast based on fungal mating peptide/G-protein-coupled receptor (GPCR) pairs harnessed from nature. First, through genome-mining, assemble 32 functional peptide-GPCR range dose-response characteristics. Next,...
We designed and synthesized synI, which is ∼21.6% shorter than native chrI, the smallest chromosome in Saccharomyces cerevisiae. SynI was for attachment to another synthetic due concerns surrounding potential instability karyotype imbalance now attached synIII, yielding first yeast fusion chromosome. Additional chromosomes were constructed study nuclear function. ChrIII-I chrIX-III-I have twisted structures, depend on silencing protein Sir3. As a smaller chromosome, chrI also faces special...
Double-strand breaks (DSBs) occur frequently during cell growth. Due to the presence of repeated sequences in genome, repair a single DSB can result gene conversion, translocation, deletion or tandem duplication depending on mechanism and sequence chosen as partner for recombinational repair. Here, we study how yeast cells single, inducible when there are several potential donors choose from, same chromosome elsewhere genome. We systematically investigate parameters that affect choice...
DNA double-strand breaks (DSBs) and other lesions occur frequently during cell growth in meiosis. These are often repaired by homologous recombination (HR). HR may result the formation of structures called Holliday junctions (HJs), which need to be resolved allow chromosome segregation. Whereas HJs present most events meiosis, it has been proposed that vegetative cells through intermediates lacking HJs. A recent screen yeast shown HJ resolution activity for a protein Yen1, addition...
Rapid and highly efficient mating-type switching of
To expand the known spectrum of genes that maintain genome stability, we screened a recently released collection temperature sensitive (Ts) yeast mutants for chromosome instability (CIN) phenotype. Proteasome subunit represented major functional group, and subsequent analysis demonstrated an evolutionarily conserved role in CIN. Analysis individual proteasome core lid mutations showed CIN phenotype at semi-permissive is associated with failure localization to nucleus. The resultant...
Significance As the use of synthetic biology grows, there is an increasing need to ensure biocontainment both protect engineered proprietary strains and mitigate potential inadvertent or advertent release environment. Here, we screen for best-performing construct essential gene-dependent transcriptional safeguards (SGs) in yeast. We have that near-WT fitness a low escape rate, grow presence nanomolar concentration supplement. also improved our “safeguard” by promoter engineering, as well...
Abstract The Sc2.0 project is building a eukaryotic synthetic genome from scratch, incorporating thousands of designer features. A major milestone has been achieved with the assembly all individual chromosomes. Here, we describe consolidation multiple chromosomes using endoreduplication intercross to generate strain 6.5 Genome-wide chromosome conformation capture and long-read direct RNA sequencing were performed on this evaluate effects modifications, such as loxPsym site insertion, tRNA...
Abstract The widely used Streptococcus pyogenes Cas9 (SpCas9) nuclease derives its DNA targeting specificity from protein-DNA contacts with protospacer adjacent motif (PAM) sequences, in addition to base-pairing interactions between guide RNA and target DNA. Previous reports have established that the PAM of SpCas9 can be altered via positive selection procedures for directed evolution or other protein engineering strategies. Here we exploit vivo systems incorporate simultaneous negative...
Cross-species pathway transplantation enables insight into a biological process not possible through traditional approaches. We replaced the enzymes catalyzing entire Saccharomyces cerevisiae adenine de novo biosynthesis with human pathway. While 'humanized' yeast grew in absence of adenine, it did so poorly. Dissection phenotype revealed that PPAT, ortholog ADE4, showed only partial function whereas all other genes complemented fully. Suppressor analysis pathways play role de-novo...
SUMMARY As part of the Synthetic Yeast 2.0 (Sc2.0) project, we designed and synthesized synthetic chromosome I. The total length synI is ∼21.4% shorter than wild-type I, smallest in Saccharomyces cerevisiae . SynI was for attachment to another due concerns potential instability karyotype imbalance. We used a variation previously developed, robust CRISPR-Cas9 method fuse I other arms varying length: chrIXR (84kb), chrIIIR (202kb) chrIVR (1Mb). All fusion strains grew like so decided attach...
Abstract Pathway transplantation from one organism to another represents a means more complete understanding of biochemical or regulatory process. The purine biosynthesis pathway, core metabolic function, was transplanted human yeast. We replaced the entire Saccharomyces cerevisiae adenine de novo pathway with cognate components. A yeast strain “humanized” for full by deleting all relevant genes completely and then providing in trans using neochromosome expressing protein coding regions...
Copyright information: Taken from Evidence for rolling circle replication of tandem genes in Nucleic Acids Research 2005;33(14):4519-4526.Published online 9 Aug 2005PMCID:PMC1184221.© The Author 2005. Published by Oxford University Press. All rights reserved DNA mutant / flies () and their heterozygous sibling flies, which are a mixture /Cy /SM1 labeled /+ was analyzed on 2D gel probed with histone H3. Arrows indicate RCIs. samples both A B contain similar levels linear as quantified...