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Authors: IGI
Group: CornLab
Proper citation: IGI 2015. Re-amplification of CRISPRa and CRISPRi libraries (v1.0)Reference: Genome-Scale CRISPR-Mediated Control of Gene Repression and A. protocols.io dx.doi.org/10.17504/protocols.io.dmz475 Copy
Authors: Breton Hornblower
Summary: T7 Endonuclease I recognizes and cleaves non-perfectly matched DNA. This protocol describes how to determine genome targeting efficiency by digesting annealed PCR products with T7 Endonuclease I. In the first step PCR products are produced from the genomic DNA of cells whose genomes were targeted using Cas9, TALEN, ZFN etc. In the second step, the PCR products are annealed and digested with T7 Endonuclease I. Fragments are analyzed to determine the efficiency of genome targeting.
Proper citation: Breton Hornblower 2016. Determining Genome Targeting Efficiency using T7 Endonuclease I (M0302). protocols.io dx.doi.org/10.17504/protocols.io.eqxbdxn Copy
Authors: Alan Cone
Group: Ju Lab
Summary: Cleans and concentrates any DNA. Typically useful after PCR, restriction digest, or anything where DNA was altered and now you need your newly altered DNA in a pure form.
Proper citation: Alan Cone 2016. Zymo Research DNA Clean & Concentrator. protocols.io dx.doi.org/10.17504/protocols.io.egbbbsn Copy
Authors: Jason Kim
Group: Mouse Metabolic Phenotyping Centers
Summary:
This experiment involves a spectrophotometric measurement using Roche Cobas Clinical Chemistry Analyzer. Serum levels of creatinine reflect systemic protein metabolism.
Proper citation: Jason Kim 2019. U Mass - Creatinine. protocols.io dx.doi.org/10.17504/protocols.io.xvjfn4n Copy
Authors: Nus Igem
Proper citation: Nus Igem 2019. Preparation of M9 Media. protocols.io dx.doi.org/10.17504/protocols.io.8cjhsun Copy
Authors: Plínio Luna de Albuquerque
Summary: It is a crossover, double-blind, sham-controlled, pseudorandomized, and counterbalanced study at the Laboratory of Applied Neuroscience (LANA) of the Federal University of Pernambuco (UFPE). We performed a double-blind, randomized, sham-controlled crossover study with 12 healthy volunteers who underwent single sessions of rTMS (1Hz, 20Hz and Sham) and tsDCS (anodal, cathodal and Sham) associated with 20 minutes of treadmill walking. Cortical excitability was assessed by motor evoked potential (MEP) and spinal cord excitability by the Hoffmann reflex (Hr), nociceptive flexion reflex (NFR) and homosynaptic depression (HD). All measures were assessed before, immediately, 30 and 60 minutes after the experimental procedures.
Proper citation: Plínio Luna de Albuquerque 2018. Effect of different types of non-invasive central nervous system stimulation on pain perception, cortical and spinal cord excitability in healthy individuals.. protocols.io dx.doi.org/10.17504/protocols.io.nkidcue Copy
Authors: Juan Carlos Herguera-García, Gisela Heckel
Summary: In the Stable Isotope Laboratory at CICESE (Ensenada Center for Scientific Research and Graduate Education, Mexico) animal tissues (pup fur from Pacific harbor seals, Phoca vitulina richardii) were ground for isotopic determinations. Samples were stored in a dissecator until they were loaded into a Costech® Carrousel. Samples were combusted at high temperature (1000˚C) in a pure oxygen atmosphere in an elemental analyzer coupled to a Finnigan MAT Delta V Advantage continuous flow stable isotope mass spectrometer.Stable carbon and nitrogen ratios are expressed as δ13C or δ15N according to the following equation:δ13C or δ15N (‰) = [Rsample/Rstandard)–1]* 1000 (1) where R is 13C/ 12C or 15N/ 14N. Isotopic values of carbon and nitrogen are reported relative to Pee Dee Belemnite and atmospheric nitrogen standards, respectively. The accuracy of isotopic measurements was verified using secondary standard reference materials (Glutamic acid, a pure carbonate Merck and the laboratory internal references Lanugo for N, and CH94 for C).
Proper citation: Juan Carlos Herguera-García, Gisela Heckel 2019. Carbon and nitrogen stable isotope analysis in harbor seal pup fur at CICESE. protocols.io dx.doi.org/10.17504/protocols.io.2m3gc8n Copy
Authors: Marie-Ka Tilak
Summary: This protocol was developed with the aim of sequencing the GC rich part of heterogeneous genomes with the Illumina technology. To this aim, fragmented genomic DNA is denaturized up to 90°C before library preparation. This procedure enriches the library in GC-rich fragments and leads to a substantial increase in average GC content of sequence reads.
Proper citation: Marie-Ka Tilak 2017. Heat fragmented genomic DNA before Illumina library preparation for sequencing GC rich fraction of heterogeneous genome . protocols.io dx.doi.org/10.17504/protocols.io.jxicpke Copy
Authors: Magdalena Julkowska
Group: Salt Lab KAUST
Summary: Standard protocol for Agrobacterium transformation
Proper citation: Magdalena Julkowska 2019. Electrocompetent Agrobacterium transformation. protocols.io dx.doi.org/10.17504/protocols.io.pewdjfe Copy
Authors: James Thornton
Group: ECOGEO
Summary: Short instructions on how to allow copying and pasting to the virtual machine.
Proper citation: James Thornton 2016. How to copy from host to guest machine. protocols.io dx.doi.org/10.17504/protocols.io.ferbjd6 Copy
Authors: Sandra Rieger
Group: Diabetic Complications Consortium
Summary: To assess hyperglycemia in larval zebrafish, we analyzed the expression of genes implicated in glucose metabolism, such as insulin, insulin receptor, glucagon, and phosphoenolpyruvate carboxylase (pepck), using quantitative PCR (qPCR). We analyzed gene expression following treatment of Tg(ins:NTRmCherry) transgenic larvae either with 0.5 % DMSO (controls) or 10 mM metronidazole (see Protocol 2) for β-cell ablation at 3 and 8 days post fertilization (dpf). Diabetic Complication:
Proper citation: Sandra Rieger 2019. Quantitative PCR analysis to assess gene expression changes in hyperglycemic larval zebrafish. protocols.io dx.doi.org/10.17504/protocols.io.3bagiie Copy
Authors: Shaina Robbins, Alison Moss, Sean Nieves
Group: SPARC
Summary: This protocol utilizes Fluidigm's Biomark system which performs high-throughput real-time PCR that can assay 48 or 96 genes for 48 or 96 samples respectfully. This protocol is used for gene expression targeting samples at the single-cell scale and can be used with the 48.48 Dynamic Array integrated fluidic circuit (IFC) or the 96.96 Dynamic Array IFC. Note that this protocol assumes that single-cell samples are captured using the Arcturus Laser Capture Microdissection system.
Proper citation: Shaina Robbins, Alison Moss, Sean Nieves 2020. BioMark Single Cell Protocol (Two-Step RTSTA). protocols.io dx.doi.org/10.17504/protocols.io.wdsfa6e Copy
Authors: Elin Einarsson
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: We have created plasmids for transfection of the parasite Perkinsus marinus using Golden Gate cloning. We have explored the possibility to either FACS sort (GFP/mCherry fused to MOE membrane protein) or using drug selection (bleomycin, blasticidin S or puromycin) to enrich transfected cells.
Proper citation: Elin Einarsson 2019. Golden Gate plasmids used for transfection of Perkinsus marinus. protocols.io dx.doi.org/10.17504/protocols.io.37egrje Copy
Authors: Jun Minagawa
Proper citation: Jun Minagawa 2019. DNA construct for genetic transformation of the coral symbiotic alga Breviolum sp.. protocols.io dx.doi.org/10.17504/protocols.io.7udhns6 Copy
Authors: Larry Klobutcher
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: Larry Klobutcher 2017. Growth of Dunaliella salina on artificial seawater.. protocols.io dx.doi.org/10.17504/protocols.io.gwpbxdn Copy
Authors: Patrick Tso, Dana Lee
Group: Mouse Metabolic Phenotyping Centers
Summary: Determinations of triglycerides in plasma/serum/lymph will be made using a Randox Triglycerides colorimetric kit. The triglycerides are determined after enzymatic hydrolysis with lipases.
Proper citation: Patrick Tso, Dana Lee 2019. U Cinn - Triglyceride Assay. protocols.io dx.doi.org/10.17504/protocols.io.xm4fk8w Copy
Authors: Eftychis Frangedakis, Marta Tomaselli, Susana Sauret-Gueto
Group: OpenPlant Project
Summary: This protocol explains how to design and clone the guide RNA target sequence into a L1 plasmid ready to accept the gRNA by cloning with BbsI. L1 plasmids are L1_lacZgRNA-Ck2 and L1_lacZgRNA-Ck3.If one gRNA target sequence is cloned into the Ck2 plasmid and another one into Ck3 one, the two L1_gRNA transcription units can be combined with an antibiotic resistance transcription unit and a MpEF1α:Cas9 transcription unit via L2 SapI Loop assembly. This allows for dual gRNA editing.
Proper citation: Eftychis Frangedakis, Marta Tomaselli, Susana Sauret-Gueto 2019. gRNA design and cloning with BbsI into Loop plasmid L1_lacZgRNA-Ck2/3. protocols.io dx.doi.org/10.17504/protocols.io.94ah8se Copy
Authors: Matthew Sullivan
Group: VERVE Net, Sullivan Lab
Proper citation: Matthew Sullivan 2016. Plating Prochlorococcus and Synechococcus strains in top agarose for plaque assays. protocols.io dx.doi.org/10.17504/protocols.io.c3vyn5 Copy
Authors: Deborah A. Howatt, Anju Balakrishnan, Hong Lu
Group: Sangderk Lee lab
Proper citation: Deborah A. Howatt, Anju Balakrishnan, Hong Lu 2017. Blood Pressure Measurement: Kent Coda 8 (Tail-Cuff). protocols.io dx.doi.org/10.17504/protocols.io.iygcftw Copy
Authors: Rebecca Hufft, Christina Alba, Amy Sahud
Summary: This protocol outlines the basic methods for measuring understory percent cover, soil moisture, tree canopy cover, and plant species richness, as well as for collecting plant specimens to be vouchered in the herbarium. These measurements estimate the abundance and presence of plant species in an area, and link them to environmental conditions (soil water and light availability) that may affect their distributions. Together, these data can reveal patterns in plant community or ecosystem processes.The understory percent cover measurements will be made using the line-point intercept method. In this method, a 25m transect is placed along the ground with a 25m by 1m belt transect to either side of it. Percent cover is measured along the transect, along with soil moisture and tree canopy cover. Plant richness is measured in the belt transect and the plant vouchering is done outside of the transect and belt transect area.
Proper citation: Rebecca Hufft, Christina Alba, Amy Sahud 2020. Vegetation Monitoring Protocol for measuring and collecting ecological data. protocols.io dx.doi.org/10.17504/protocols.io.bkfuktnw Copy
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