- PA5-87292 - Invitrogen Antibodies QKI antibody

Submitted references Actin remodeling driven by circLIMA1: sperm cell as an intriguing cellular model.

Manfrevola F, Potenza N, Chioccarelli T, Di Palo A, Siniscalchi C, Porreca V, Scialla A, Mele VG, Petito G, Russo A, Lanni A, Senese R, Ricci G, Pierantoni R, Chianese R, Cobellis G
International journal of biological sciences 2022;18(13):5136-5153

FUS driven circCNOT6L biogenesis in mouse and human spermatozoa supports zygote development.

Chioccarelli T, Falco G, Cappetta D, De Angelis A, Roberto L, Addeo M, Ragusa M, Barbagallo D, Berrino L, Purrello M, Ambrosino C, Cobellis G, Pierantoni R, Chianese R, Manfrevola F
Cellular and molecular life sciences : CMLS 2021 Dec 22;79(1):50

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of QKI in MCF-7 cells. Sample was incubated with polyclonal QKI antibody (Product # PA5-87292).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 4 FUS drives CNOT6L backsplicing by interacting with RNApol2 and QKI in caput SPZ. The enrichment levels of circCNOT6L and CNOT6L-mRNA in the products of RIP assay (FUS-IP compared with IgG-IP) in WT caput SPZ alone ( A ) and after in vitro ACEA treatment ( F ) detected by qRT-PCR. Data are reported as mean +- SEM from three independent experiments. ** P < 0.01. qRT-PCR detection of circCNOT6L and CNOT6L-mRNA expression levels ( E ) in caput SPZ from WT mice in vitro treated with vehicle (CTRL) or ACEA 1 uM ( n = 3 different samples from three different animals for each experimental group in triplicate). Data are reported as mean value of nfe +- SEM, using Actin as endogenous control. ** P < 0.01. Western blot analysis of FUS, QKI and RNApol2 in the products of IP in WT caput SPZ ( B , C and D ) using FUS, QKI and RNApol2 antibodies. Western blot analysis of RIP protein fraction immunoprecipitated with FUS Ab (FUS-IP) in WT caput SPZ after in vitro ACEA treatment ( G ). FUS-IP is analyzed in comparison to control IgG-IP and Input protein extracts. qRT-PCR detection of Fus expression levels in ESCs, cultured in RM medium, treated with siScr or siFus (5, 10, 25 and 50 nM) and harvested after 48 ( H ) and 72 h ( I ) from siRNA transfection. Data are reported as mean value of nfe +- SEM, using Actin as endogenous control. ** P < 0.01; * P < 0.05. Western blot analysis of FUS protein ( J ) and qRT-PCR detection of circCNOT6L expression levels ( K ) in ESCs, cultured in RM med

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 7 Human SPZ mimic mouse SPZ. qRT-PCR detection of circCNOT6L and CNOT6L-mRNA expression levels ( A ); immunoblots and quantification of CB1 ( B ), FUS ( C ) and ADAR ( D ) proteins in A- and B-SPZ fractions from normozoospermic volunteers ( n = 5 different samples in triplicate). Immunofluorescence analysis of FUS protein in A- and B-SPZ ( E and F ). White empty arrowheads and white full arrowheads represent FUS localization (FITC-green) in sperm head and tail, respectively. Nuclei were labeled with DAPI (blue). Scale bar: 20 uM. qRT-PCR detection of circCNOT6L and CNOT6L-mRNA expression levels ( G ) in B-SPZ fractions from normozoospermic volunteers ( n = 5) in vitro treated with vehicle (CTRL) or ACEA 1 uM ( n = 5 different samples for each experimental group in triplicate). In ( A ) and ( G ), the data are reported as mean value of nfe +- SEM, using Gapdh as endogenous control. In ( B ), ( C ) and ( D ) CB1, FUS and ADAR amount was quantified by densitometry analysis, normalized against Tubulin (TUB) signals, expressed as fc of OD values and reported as mean value +- SEM. ** P < 0.01. The enrichment levels of circCNOT6L and CNOT6L-mRNA in the products of RIP assay (FUS-IP compared with IgG-IP) in B-SPZ alone ( H ) and after in vitro ACEA treatment ( L ) by qRT-PCR. Data are reported as mean +- SEM from three independent experiments. ** P < 0.01. Western blot analysis of FUS, QKI and RNApol2 in the products of IP in B-SPZ ( I , J and K ) using FUS, QKI and RNApol2 anti

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: CircLIMA1-dependent sperm nuclear actin organization along the epididymis. qRT-PCR detection of circLIMA1 expression levels in caput and cauda SPZ ( A ) or epididymis ( B ) from WT and CB1 -/- mice. ( C ) Immunofluorescence analysis of F-actin by phalloidin staining in caput and cauda SPZ from WT and CB1 -/- mice. qRT-PCR detection of circLIMA1 expression levels ( D ) and Immunofluorescence analysis of F-actin by phalloidin staining ( E ) in caput SPZ from WT mice in vitro co-incubated with: WT Caput ELF (CTRL group), WT Caput ELF combined with Cytochalasin-D (C8273; Sigma-Aldrich, Milano, Italy) at the concentration of 10 muM (ELF WT+CYTOD); CB1 -/- Caput ELF (ELF CB1 -/- ); CB1 -/- Caput ELF combined with Cytochalasin-D 10 muM (ELF CB1 -/- +CYTOD); (n=3 different samples for each experimental group from 8 different animals in triplicate). qRT-PCR data are normalized using Cyclophilin and RPS18 , for SPZ and epididymis respectively, expressed as DeltaCq and reported as mean value +- S.E.M. Experimental groups with statistically significant differences (p

PA5-87292

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