Takara Bio Cellartis iPSC rCas9, Руководство пользователя

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Cellartis iPSC rCas9 Electroporation and Single-Cell Cloning System User Manual
(030619)
takarabio.com
Takara Bio USA, Inc.
Page 7 of 24
III. Additional Materials Not Supplied
A. Required
Mammalian Cell Culture Supplies
Use the Cellartis DEF-CS 500 Culture System (Takara Bio, Cat. No. Y30010) for maintaining hiPS cell
lines 1) prior to using the Cellartis iPSC rCas9 Electroporation and Single-Cell Cloning System and 2) for
scaling up clonal lines created using this system.
The following tissue culture materials are required but not supplied:
•
96-well plates, flat bottom, cell-culture treated (Corning, Cat. No. 3595 or equivalent)
•
48-well plates, flat bottom, cell-culture treated (Corning, Cat. No. 3548 or equivalent)
•
TrypLE Select Enzyme (1X), no phenol red (Thermo Fisher Scientific, Cat. No. 12563011)
•
PBS Dulbecco's with Ca
2+
& Mg
2+
(D-PBS +/+) (Sigma, Cat. No. D8662 or equivalent)
•
PBS Dulbecco's w/o Ca
2+
& Mg
2+
(D-PBS –/–) (Sigma, Cat. No. D8537 or equivalent)
General Supplies
•
96–100% ethanol (Sigma, Cat. No. E7023 or equivalent)
•
Isopropanol (Sigma, Cat. No. I9516 or equivalent)
•
NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Cat. No. ND-2000 or equivalent)
Electroporation Supplies
Use of this product requires an electroporator, electroporation chamber (typically cuvettes or tips), and
an electroporation buffer that is suitable for hiPSCs. In this manual, we provide a protocol for
electroporation using the Neon Transfection System (Thermo Fisher Scientific, Cat. No. MPK5000).
If using the 4D-Nucleofector System (Lonza, Cat. No. AAF-1002B), please refer to the
manufacturer’s website for operating instructions.
B. Recommended
sgRNA Efficacy Testing Supplies
If you to wish to test the efficacy of different sgRNAs in vitro prior to using them in editing experiments,
try the Guide-it sgRNA Screening Kit (Cat. No. 632639).
Genotype Confirmation Supplies
These items are recommended for determining the efficiency of gene editing after electroporation (Cat.
Nos. 631443 & 631448), the genotype (Cat. No. 632611), and the sequence of the edits (Cat. No. 631444)
in the clonal cell lines:
Cat. No. Product Size
631443 Guide-it Mutation Detection Kit 100 rxns
631448 Guide-it Mutation Detection Kit 25 rxns
632611 Guide-it Genotype Confirmation Kit 100 rxns
631444 Guide-it Indel Identification Kit 10 rxns