Anti-DNA ligase1 (Human) antibody, rabbit polyclonal
Product code70-090Size

100 µgStorage-20℃

Concentration

1.0 mg/ml

Buffer

PBS- with 50% glycerol

PurityPurified IgG fraction with protein A from rabbit antiserum.

Immunogen

Recombinant FLAG-Lig1 highly purified from insect cells infected withbaculovirus designed for expression of the full-length FLAG-Lig1.

IsotypeRabbit IgG

Reactivity

Human cell lysatesSpecial notes

N/A

Application

1. Western blotting: 1 µg/ml (Fig.1)
2. Immunofluorescent staining: 0.5 µg/ml (Fig.2)

Background

LIG1 (HGNC: 6598) encodes one of the ATP-dependent DNA ligases in humancells, which functions in DNA replication, recombination, and the base excision
repair process. Mutations in this gene lead to DNA ligase I deficiency and resultin immunodeficiency and increased sensitivity to DNA-damaging agents,suggesting its relation to cancer development. Human cells have two additional
DNA ligase families, LIG3 and LIG4, both of which have redundant roles toLIG1 in DNA replication and DNA damage responses and their mutations arealso involved in cancer development (Annu Rev Biochem. 2008; 77: 313–338).
Several Lig1-interacting proteins have been identified. One of the interacting
proteins is DNA-sliding clamp, the homotrimeric proliferating cell nuclear
antigen (PCNA), which are involved in DNA replication and repair. Theinteraction with PCNA is primarily mediated by a conserved PCNA-binding
motif, PIP box, at the N terminus of Lig1. The interaction with PCNA is required
for localization of Lig1 at the sites of DNA replication, the Okazaki fragmentjoining and the base excision repair.

Data LinkUniProtKB - P18858 (DNLI1_HUMAN)

Please note: All products are FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC
PROCEDURES. NOT FOR MILITARY USE.
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Data Images: 70-090 Anti-DNA ligase1 (Human) antibody, rabbit polyclonal

Fig. 1 Detection of endogenous Lig1 in whole cell extract of human HEK293T cells
34 μg of the HEK293T cell lysate was separated on 10% SDS-PAGE and blotted on PVDF membranewith a wet-transfer apparatus at 15V overnight. After blocking of the membrane with 5% skim milkfor 95min, anti-Lig1 antibody was applied to it at 1μg/ml for 145min, and then, Goat Anti-Rabbit IgG
H&L (HRP) (ab205718) was applied at 1/10,000 dilution at RT for 90min. The reacted antibody wasdetected with Immunostar zeta reagent (291-72401, wako). A 140kDa band corresponding to thecellular Lig1 could be detected.

Fig. 2 Immunofluorescence detection of HeLa cell nuclei
Hela cell grown in a dish was fixed with 4 % PFA at 4℃ for 30min. and permeabilized with 0.25 %Triton X-100 in PBS- for 10 min. 0.5μg/ml of anti-RPA antibody was applied to the sample at 4℃
overnight after blocking with 2% BSA in PSB- at RT for 75min. The reacted antibody was detectedwith Goat anti rabbit IgG-H&L (Alexa Fluor 488) (ab150077) at 1/1000 dilution for 70min. DAPI
staining was done with 1.0μg/mL DAPI in TBS for 5 min. Fluorescent signals were observed by using
Ts2-FL (Nicon). Fluorescent images were analyzed in NIS-Elements D (Nicon).
The nuclei were stained specifically with this antibody.
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Fig.3 Inhibition of ligase activity by anti Lig-1 antibody
1.λHind III digest + Ligase-1
2.λHind III digest + Ligase-1 + + anti Lig-1 antibody
3.λHind III digest
protocol:Ligase-1 + anti Lig-1 antibody reaction on ice for 30min
add λHind III digest at 16℃ for 30min
Reference:
1. Patrick Maffucci, et al. Biallelic mutations in DNA ligase 1 underlie a spectrum of immune
deficiencies. J.Clin Invest. 128(12). :5489-5504 (2018) PMID: 30395541. WB

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