Restriction enzymes (BamHI and EcoRI) and T4 ligase were purchased from Sangon Biotech Co., Ltd. Rabbit polyclonal or monoclonal antibodies against human cytokeratin 7 (cat. no. ab181598), Bax (cat. no. ab32503), Bcl-2 (cat. no. ab196495), PI3K (cat. no. ab86714), optineurin (OPTN; cat. no. ab151240), Akt1/2/3 (cat. no. ab184136), phosphorylated (p)-Akt1 (T308; cat. no. ab105731), excision repair cross-complementation group 1 (ERCC1; cat. no. ab129267) and β-actin (cat. no. ab8227) were purchased from Abcam. Rabbit polyclonal antibodies against human caspase-3 p17 (cat. no. sc-271,028) and EEF1D (cat. no. abs136494) were purchased from Santa Cruz Biotechnology, Inc., and Absin Bioscience, Inc., respectively. Lipofectamine® 3000 transfection reagent was purchased from Invitrogen (Thermo Fisher Scientific, Inc.). The HRP-conjugated secondary antibody (goat anti-mouse or goat anti-rabbit IgG, cat. no. PV-6000) were purchased from Beijing Zhongshan Jinqiao Biotechnology Co., Ltd. FITC-labeled goat anti-rabbit IgG (H + L) secondary antibody (cat. no. A0562) and Hoechst 33342 (cat. no. C1022) were purchased from Beyotime Institute of Biotechnology. LY294002 (PI3K inhibitor; cat. no. ab120243) and MK-2206 (Akt inhibitor; cat. no. SF2712) were purchased from Abcam and Beyotime Institute of Biotechnology, respectively.
T4 ligase
Manufactured by Sangon
T4 ligase is a DNA ligase enzyme derived from the T4 bacteriophage. Its primary function is to catalyze the formation of phosphodiester bonds between adjacent 3'-hydroxyl and 5'-phosphate ends in double-stranded DNA molecules, allowing for the joining of DNA fragments.
Automatically generated - may contain errors
2 protocols using t4 ligase
1
Antibody and Reagents for Cell Signaling
2
Construction of Cloning Vectors with Antibiotic Resistance
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The oligonucleotide primers used for the construction of the cloning vector are listed in Table 2. Plasmid pUC19 was double-digested with SacI and BamHI. The Em resistance gene was amplified by PCR using the primers EmF and EmR, and pMG36e as the template. Then, DNA fragments of different lengths were produced by PCR amplification using pLC2W DNA as the template. All DNA fragments were ligated by T4 Ligase (Sangon Biotech). Plasmids were constructed using different lengths of the pLC2W replicon.
The Em resistance gene of pUE-F1 was replaced in a 2-step process by a chloramphenicol (Cm) resistance gene from pNZ5319, giving rise to pUCM-F1. The availability of constructs with different antibiotic selections allowed the consecutive transformation of pUE-F1 and pUCM-F1 into LAB.
The Em resistance gene of pUE-F1 was replaced in a 2-step process by a chloramphenicol (Cm) resistance gene from pNZ5319, giving rise to pUCM-F1. The availability of constructs with different antibiotic selections allowed the consecutive transformation of pUE-F1 and pUCM-F1 into LAB.
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