Thalassiosira spp. Community Composition Shifts in Response to Chemical and Physical Forcing in the Northeast Pacific Ocean

Diatoms are genetically diverse unicellular photosynthetic eukaryotes that are key primary producers in the ocean. Many of the over 100 extant diatom species in the cosmopolitan genus Thalassiosira are difficult to distinguish in mixed populations using light microscopy. Here, we examine shifts in Thalassiosira spp. composition along a coastal to open ocean transect that encountered a 3-month-old Haida eddy in the northeast Pacific Ocean. To quantify shifts in Thalassiosira species composition, we developed a targeted automated ribosomal intergenic spacer analysis (ARISA) method to identify Thalassiosira spp. in environmental samples. As many specific fragment lengths are indicative of individual Thalassiosira spp., the ARISA method is a useful screening tool to identify changes in the relative abundance and distribution of specific species. The method also enabled us to assess changes in Thalassiosira community composition in response to chemical and physical forcing. Thalassiosira spp. community composition in the core of a 3-month-old Haida eddy remained largely (\u3e80%) similar over a 2-week period, despite moving 24 km southwestward. Shifts in Thalassiosira species correlated with changes in dissolved iron (Fe) and temperature throughout the sampling period. Simultaneously tracking community composition and relative abundance of Thalassiosira species within the physical and chemical context they occurred allowed us to identify quantitative linkages between environmental conditions and community response

\u3cem\u3eThalassiosira spp.\u3c/em\u3e Community Composition Shifts in Response to Chemical and Physical Forcing in the Northeast Pacific Ocean

Diatoms are genetically diverse unicellular photosynthetic eukaryotes that are key primary producers in the ocean. Many of the over 100 extant diatom species in the cosmopolitan genus Thalassiosira are difficult to distinguish in mixed populations using light microscopy. Here, we examine shifts in Thalassiosira spp. composition along a coastal to open ocean transect that encountered a 3-month-old Haida eddy in the northeast Pacific Ocean. To quantify shifts in Thalassiosira species composition, we developed a targeted automated ribosomal intergenic spacer analysis (ARISA) method to identify Thalassiosira spp. in environmental samples. As many specific fragment lengths are indicative of individual Thalassiosira spp., the ARISA method is a useful screening tool to identify changes in the relative abundance and distribution of specific species. The method also enabled us to assess changes in Thalassiosira community composition in response to chemical and physical forcing. Thalassiosira spp. community composition in the core of a 3-month-old Haida eddy remained largely (\u3e80%) similar over a 2-week period, despite moving 24 km southwestward. Shifts in Thalassiosira species correlated with changes in dissolved iron (Fe) and temperature throughout the sampling period. Simultaneously tracking community composition and relative abundance of Thalassiosira species within the physical and chemical context they occurred allowed us to identify quantitative linkages between environmental conditions and community response

Mouse Intestine Selects Nonmotile \u3cem\u3eflhDC\u3c/em\u3e Mutants of \u3cem\u3eEscherichia coli\u3c/em\u3e MG1655 with Increased Colonizing Ability and Better Utilization of Carbon Sources

d-Gluconate which is primarily catabolized via the Entner-Doudoroff (ED) pathway, has been implicated as being important for colonization of the streptomycin-treated mouse large intestine by Escherichia coli MG1655, a human commensal strain. In the present study, we report that an MG1655 Δedd mutant defective in the ED pathway grows poorly not only on gluconate as a sole carbon source but on a number of other sugars previously implicated as being important for colonization, including l-fucose, d-gluconate, d-glucuronate, N-acetyl-d-glucosamine, d-mannose, and d-ribose. Furthermore, we show that the mouse intestine selects mutants of MG1655 Δedd and wild-type MG1655 that have improved mouse intestine-colonizing ability and grow 15 to 30% faster on the aforementioned sugars. The mutants of MG1655 Δedd and wild-type MG1655 selected by the intestine are shown to be nonmotile and to have deletions in the flhDC operon, which encodes the master regulator of flagellar biosynthesis. Finally, we show that ΔflhDC mutants of wild-type MG1655 and MG1655 Δedd constructed in the laboratory act identically to those selected by the intestine; i.e., they grow better than their respective parents on sugars as sole carbon sources and are better colonizers of the mouse intestine

BBF RFC 94: Type IIS Assembly for Bacterial Transcriptional Units: A Standardized Assembly Method for Building Bacterial Transcriptional Units Using the Type IIS Restriction Enzymes BsaI and BbsI

This RFC94 describes an assembly standard based on the Type IIS restriction enzymes BsaI and BbsI (also called BpiI). This assembly standard is based upon the Modular Cloning (MoClo) assembly strategy, which was introduced in 2011 by Weber et al. [1] and is based upon Golden Gate cloning [2]. In this RFC, we describe our proposed MoClo standard for generating a library of bacterial DNA parts for generating four-part transcriptional units (promoter : 5’UTR : CDS : 3’UTR). In this work, we define 5’UTRs as including ribosomal binding sites (RBS) and bi-cistronic design elements (BCDs) [3], and 3’UTRs as transcriptional terminators. The 2012-2014 BostonU iGEM teams completed this work and a more compact library has also been created based on this work [4]

CIDAR MoClo: Improved MoClo Assembly Standard and New E. coli Part Library Enable Rapid Combinatorial Design for Synthetic and Traditional Biology

Multipart and modular DNA part libraries and assembly standards have become common tools in synthetic biology since the publication of the Gibson and Golden Gate assembly methods, yet no multipart modular library exists for use in bacterial systems. Building upon the existing MoClo assembly framework, we have developed a publicly available collection of modular DNA parts and enhanced MoClo protocols to enable rapid one-pot, multipart assembly, combinatorial design, and expression tuning in Escherichia coli. The Cross-disciplinary Integration of Design Automation Research lab (CIDAR) MoClo Library is openly available and contains promoters, ribosomal binding sites, coding sequence, terminators, vectors, and a set of fluorescent control plasmids. Optimized protocols reduce reaction time and cost by >80% from that of previously published protocols

Mouse Intestine Selects Nonmotile flhDC Mutants of Escherichia coli MG1655 with Increased Colonizing Ability and Better Utilization of Carbon Sources

d-Gluconate which is primarily catabolized via the Entner-Doudoroff (ED) pathway, has been implicated as being important for colonization of the streptomycin-treated mouse large intestine by Escherichia coli MG1655, a human commensal strain. In the present study, we report that an MG1655 Δedd mutant defective in the ED pathway grows poorly not only on gluconate as a sole carbon source but on a number of other sugars previously implicated as being important for colonization, including l-fucose, d-gluconate, d-glucuronate, N-acetyl-d-glucosamine, d-mannose, and d-ribose. Furthermore, we show that the mouse intestine selects mutants of MG1655 Δedd and wild-type MG1655 that have improved mouse intestine-colonizing ability and grow 15 to 30% faster on the aforementioned sugars. The mutants of MG1655 Δedd and wild-type MG1655 selected by the intestine are shown to be nonmotile and to have deletions in the flhDC operon, which encodes the master regulator of flagellar biosynthesis. Finally, we show that ΔflhDC mutants of wild-type MG1655 and MG1655 Δedd constructed in the laboratory act identically to those selected by the intestine; i.e., they grow better than their respective parents on sugars as sole carbon sources and are better colonizers of the mouse intestine

<i>Owl</i>: Electronic Datasheet Generator

<i>Owl</i> (www.owlcad.org) is a biodesign automation tool that generates electronic datasheets for synthetic biological parts using common formatting. Data can be retrieved automatically from existing repositories and modified in the <i>Owl</i> user interface (UI). <i>Owl</i> uses the data to generate an HTML page with standard typesetting that can be saved as a PDF file. Here we present the <i>Owl</i> software tool in its alpha version, its current UI, its description of input data for generating a datasheet, its example datasheets, and the vision of the tool’s role in biodesign automation