Journal: Nature Protocols

Article Title: Large-scale reconstruction of cell lineages using single-cell readout of transcriptomes and CRISPR–Cas9 barcodes by scGESTALT

doi: 10.1038/s41596-018-0058-x

Figure Lengend Snippet: Fig. 4 | Barcode copy-number determination. Copy number of scGESTALT barcode integration was determined using qPCR. Samples 1–13 represent different genomic DNA samples (n = 13 animals). Reference standards: c1 is a negative control (n = 1 animal); c2 is known to contain one copy (n = 1 animal); and c3 is known to contain two copies (n = 1 animal). This procedure was approved by the HU/FAS Committee on the Use of Animals in Research & Teaching under protocol no. 25–08.

Article Snippet: ● 35-μm Cell strainer (VWR, cat. no. 352235) ● 20-μm Cell strainer (Sysmex, cat. no. 04-004-2325) ● 1.5-ml Microcentrifuge tubes (USA Scientific, cat. no. 1415-2600) ● 1.5-ml Microcentrifuge tubes (USA Scientific, cat. no. 1415-2500) ● 0.2-ml PCR tubes (VWR, cat. no. 53509-304) ● 50-ml Tubes (VWR, cat. no. 352070) ● 15-ml Tubes (VWR, cat. no. 352096) ● 5-ml Sterile serological pipettes (VWR, cat. no. 89130-896) ● NanoDrop spectrophotometer (Thermo Scientific, cat. no. ND-2000) ● PCR and qPCR machines (Bio-Rad, model no. C-1000 and CFX96 Touch) ● DNA electrophoresis system (Bio-Rad, cat. no. 1640301) ● Benchtop centrifuge for 1.5-ml tubes (Eppendorf, cat. no. 54124) ● Refrigerated centrifuge for 1.5-ml tubes (Eppendorf, cat. no. 5424R) ● Minifuge (Benchmark, cat. no. C1008-C) ● Water bath (VWR, cat. no. 89501-460) ● Heat block for 1.5-ml tubes (VWR, cat. no. 10153-348) ● Vortex (Scientific Industries, cat. no. SI-0236) ● Magnetic rack for 0.2-ml PCR strips (Permagen, cat. no. MSR1224) ● 2100 Electrophoresis Bioanalyzer instrument (Agilent, cat. no. G2939AA) ● Laser-based micropipette puller (Sutter Instrument, cat. no. P-2000) ● Microinjection glass capillaries (World Precision Instruments, cat. no. TW100F-4) ● Fine forceps (Fine Science Tools, cat. no. 11254-20) ● Fine scissors (Fine Science Tools, cat. no. 14040-10) ● Microloader tips for glass capillaries (Eppendorf, cat. no. 930001007) ● Injection molds (Adaptive Science Tools, model no. TU-1) ● Pneumatic PicoPump (World Precision Instruments, model no. PV820) ● Piconozzle Kit v2 (World Precision Instruments, cat. no. 5430-ALL) ● Manipulator, magnetic stand, and iron plate (Narishige, model. no. M-152, GJ-1, IP) ● Treadlite II Foot Switch (Linemaster, model no. T-91-S) ● Micrometer for calibration (AmScope, cat. no. MR100) ● Dissection microscope with light source (Zeiss, model no. Stemi 2000) ● Fluorescence microscope with GFP filter (Zeiss) ● 10-cm Petri dishes (VWR, cat. no. 25384-342) ● Insect pins (Fine Science Tools, cat. no. 26002-15) ● Breeding tanks and 0.5-, 1-, and 2-L tanks (Techniplast, custom order) ● 95% O2:5% CO2 gas tank (Airgas, cat. no. X02OX95C2003102) ● Gas regulator (VWR, cat. no. 55850-392) ● Tygon tubing for bubbling gas (Component Supply, cat. no. E-3603) ● Hemocytometer (Bulldog Bio, cat. no. DHCN420) ● Qubit fluorometer (Thermo Fisher Scientific, cat. no. Q33226) ● Qubit assay tubes (Thermo Fisher Scientific, cat. no. Q32856) PROTOCOL NATURE PROTOCOLS 10 NATURE PROTOCOLS |www.nature.com/nprot

Techniques: Negative Control