Journal: Cell

Article Title: Deconstruction of corticospinal circuits for goal-directed motor skills

doi: 10.1016/j.cell.2017.08.014

Figure Lengend Snippet: KEY RESOURCE TABLE

Article Snippet: table ft1 table-wrap mode="anchored" t5 REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Chicken monoclonal anti-GFP Abcam Cat#ab1397 0 Rabbit polyclonal anti-RFP Abcam Cat#ab3477 1 Mouse monoclonal anti-NeuN Millipore Cat#MAB37 7 Rabbit polyclonal anti-PKC gamma Santa Cruz Cat#sc211 Goat polyclonal cholinergic acetyltransferase Millipore Cat#AB144 P Biological Samples N/A N/A N/A Chemicals, Peptides, and Recombinant Proteins Diphtheria toxin Sigma Cat# D0564 Green RetrobeadsTM Lumafluor Cat#N/A Clozapine N-oxide Enzo Life Sciences Cat#BML-NS105-0025 Critical Commercial Assays N/A N/A N/A Deposited Data N/A N/A N/A Experimental Models: Cell Lines N/A N/A N/A Experimental Models: Organisms/Strains Mouse/C57Bl/6 Charles River Strain code#027 Mouse/Emx1Cre The Jackson Laboratory Jax#5628 Mouse/Rosa-LSL-ChR2EYFP The Jackson Laboratory Jax#12569 Mouse/ChAT-Cre The Jackson Laboratory Jax#028861 Mouse/Thy1-ChR2EYFP The Jackson Laboratory Jax#7612 Recombinant DNA AAV-GFP This paper Cat#N/A AAV-FLEX-DTR This paper Cat#N/A AAV-FLEX-PLAP This paper Cat#N/A AAV-syn-FLEX-GCaMP6 UPenn Cat#PV2821 AAV-syn-FLEX-hM4Di-mCitrine Addgene Cat#44362 AAV-CAG-FLEX-GFP Addgene Cat#28304 AAV-EF1α-FLEX-tdTomato Addgene Cat#50462 AAV-tdTomato-P2A-SypEGFP This paper Cat#N/A AAV-FLEX-tdTomato-P2A-SypEGFP Addgene Cat# 51509 AAV-CAG-FLEX-oG-WPRE-SV40pA Addgene Cat#74292 Lenti-HiRet-GFP This paper Cat#N/A Lenti-HiRet-mCherry This paper Cat#N/A Lenti-HiRet-Cre This paper Cat#N/A Lenti-HiRet-FLEX-DTR This paper Cat#N/A Rabies-ΔG-mCherry Kim et al., 2016 Cat#N/A Rabies-ΔG-GFP Kim et al., 2016 Cat#N/A Sequence-Based Reagents N/A N/A N/A Software and Algorithms Matlab 2016b Mathworks https://www.mathworks.com/ pClamp 10 Molecular Devices https://www.moleculardevices.com/systems/conventional-patch-clamp/pclamp-10-software ImageJ2 NIH https://imagej.nih.gov/ij/index.html ImageJ Stabilizer N/A http://www.cs.cmu.edu/~kangli/code/Image_Stabilizer.html Imaris 8.0 Bitplane http://www.bitplane.com/imaris/imaris Matlab CellSort toolbox Matlab central https://www.mathworks.com/matlabcentral/fileexchange/25405-emukamel-cellsort Matlab fastICA toolbox Matlab central https://www.mathworks.com/matlabcentral/linkexchange/links/2115-the-fastica-package-for-matlab Inscopix nVista 2.0.32 Inscopix https://www.inscopix.com/ Mosaic Inscopix https://www.inscopix.com/ Other N/A N/A N/A Open in a separate window KEY RESOURCE TABLE CONTACT FOR REAGENT AND RESOURCE SHARING Further information and requests for reagent will be addressed by the lead author Zhigang He ( ude.dravrah.snerdlihc@eH.gnagihZ ).

Techniques: Recombinant, Sequencing, Software

Journal: PloS one

Article Title: ZO-1 knockout by TALEN-mediated gene targeting in MDCK cells: involvement of ZO-1 in the regulation of cytoskeleton and cell shape.

doi: 10.1371/journal.pone.0104994

Figure Lengend Snippet: Figure 1. Construction of TALENs and ZO-1 gene knockout in MDCK I and II cells. (A) TALEN binding sites in the ZO-1 gene. The left and right arms of TALEN targeting sites are indicated in blue and the spacer region is indicated in red. The initiation codon within the spacer region is highlighted. (B) Immunofluorescence microscopic analysis of ZO-1, ZO-2 and ZO-3 in MDCK II cells transfected with TALEN constructs for ZO-1 gene knockout. After transfection, cells were subcultured on filter inserts for 4 d before analysis. At the boundary of control and ZO-1 knockout cells, characteristic convex curves of cell–cell junctions are observed (arrows). (C) Immunofluorescence microscopic analysis of ZO-1, ZO-2 and ZO-3 in MDCK I cells transfected with TALEN constructs for ZO-1 gene knockout. Similar morphological changes of cell–cell junctions at the boundary of control and ZO-1 knockout cells were observed in MDCK I cells. Staining of ZO-3 was reduced in ZO-1 knockout cells (arrowheads). Scale bars, 10 mm. doi:10.1371/journal.pone.0104994.g001

Article Snippet: Construction of TALENs and establishment of knockout clones TALENs were constructed following the detailed instruction provided by the TALE Toolbox kit from the Zhang laboratory [27] (Addgene, #1000000019).

Techniques: TALENs, Gene Knockout, Binding Assay, Immunofluorescence, Transfection, Construct, Control, Knock-Out, Staining

Journal: PloS one

Article Title: ZO-1 knockout by TALEN-mediated gene targeting in MDCK cells: involvement of ZO-1 in the regulation of cytoskeleton and cell shape.

doi: 10.1371/journal.pone.0104994

Figure Lengend Snippet: Figure 4. Establishment of ZO-1 knockout clones in MDCK II cells. (A) Immunofluorescence microscopic analysis of ZO-1 in control (CTL) MDCK II cells and ZO-1 knockout clones (KO 1–3). ZO-1 staining was completely lost in ZO-1 knockout clones. Scale bar, 10 mm. (B) Immunoblots of ZO-1 and E-cadherin (E-cad) in control MDCK II cells and ZO-1 knockout clones. Knockout clones showed no detectable bands of ZO-1. (C) DNA sequences of TALEN targeting sites in each allele of ZO-1 knockout clones. One type of mutation was present in the alleles of ZO-1 knockout clone 1 (KO 1) and two types of mutations in the alleles of clones 2 and 3 (KO 2 and 3). Dashes indicate loss of nucleotides and green letters indicate additional nucleotides. Loss of initiating codon or frameshift were confirmed in all alleles. (D) Genomic PCR analysis of control and ZO-1 knockout clones using primers for TALENs and ZO-1 DNAs. A clone stably expressing TALEN was used as a positive control (PC). None of the PCR products for TALENs were detected in ZO-1 knockout clones. doi:10.1371/journal.pone.0104994.g004

Article Snippet: Construction of TALENs and establishment of knockout clones TALENs were constructed following the detailed instruction provided by the TALE Toolbox kit from the Zhang laboratory [27] (Addgene, #1000000019).

Techniques: Knock-Out, Clone Assay, Immunofluorescence, Control, Staining, Western Blot, Mutagenesis, TALENs, Stable Transfection, Expressing, Positive Control