pcluc mini tk 2 vector  (New England Biolabs)


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    Name:
    pCLuc Basic 2 Vector
    Description:
    pCLuc Basic 2 Vector 20 ug
    Catalog Number:
    N0317S
    Price:
    307
    Category:
    Cellular Biology
    Size:
    20 ug
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    Structured Review

    New England Biolabs pcluc mini tk 2 vector
    pCLuc Basic 2 Vector
    pCLuc Basic 2 Vector 20 ug
    https://www.bioz.com/result/pcluc mini tk 2 vector/product/New England Biolabs
    Average 91 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    pcluc mini tk 2 vector - by Bioz Stars, 2021-05
    91/100 stars

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    Related Articles

    Clone Assay:

    Article Title: HNF1A is a novel oncogene that regulates human pancreatic cancer stem cell properties
    Article Snippet: For the POU5F1/OCT4 LTR construct, a 1.7-kbp insert was amplified from NY5 genomic DNA with the following primers: 5’-ATCTTGGAATTCTGGGCACTCAGTTTATTGTTAGG-3’ and 5’-GGTGGCGGATCCTGTGTTAATCCTCCTCGGGG-3’. .. The insert was digested with EcoRI and BamHI and cloned into pCLuc-Basic2. .. Cypridina luciferase constructs were co-transfected with Lipofectamine 2000 (Invitrogen) into 293FT cells with either LacZ or HNF1A lentiviral expression plasmids and the internal control plasmid pTK-GDLuc, a variant of pTK-GLuc (New England Biolabs) in which the Gaussia luciferase coding region was replaced with the coding region for Gaussia-Dura (Millipore) in order to provide a more stable luciferase signal.

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Luciferase:

    Article Title: Ribophorin II regulates breast tumor initiation and metastasis through the functional suppression of GSK3?
    Article Snippet: The PCR fragment was inserted into BglII- and SalI-treated pEGFP-1 (Clontech). .. To obtain a HSP70 promoter-driven secreted cypridina luciferase (pHSP70-CLuc), HSP70-GFP was digested with KpnI and NotI and the fragment containing GFP was replaced with cypridina luciferase gene derived from pCLuc-Basic2 (NEB). ..

    Article Title: HNF1A is a novel oncogene that regulates human pancreatic cancer stem cell properties
    Article Snippet: Biotinylated mouse anti-mouse H-2Kd/H-2Dd clone 34-1-2S was purchased from SouthernBiotech (Birmingham, AL). .. For the Cypridina luciferase construct containing the full-length canonical OCT4 promoter, a 3.9-kbp insert was excised from phOct4-EGFP ( ) by XhoI and BamHI digestion, followed by ligation into pCLuc-Basic2 (New England Biolabs). phOct4-EGFP was a gift from Wei Cui (Addgene plasmid # 38776). .. For the POU5F1/OCT4 LTR construct, a 1.7-kbp insert was amplified from NY5 genomic DNA with the following primers: 5’-ATCTTGGAATTCTGGGCACTCAGTTTATTGTTAGG-3’ and 5’-GGTGGCGGATCCTGTGTTAATCCTCCTCGGGG-3’.

    Derivative Assay:

    Article Title: Ribophorin II regulates breast tumor initiation and metastasis through the functional suppression of GSK3?
    Article Snippet: The PCR fragment was inserted into BglII- and SalI-treated pEGFP-1 (Clontech). .. To obtain a HSP70 promoter-driven secreted cypridina luciferase (pHSP70-CLuc), HSP70-GFP was digested with KpnI and NotI and the fragment containing GFP was replaced with cypridina luciferase gene derived from pCLuc-Basic2 (NEB). ..

    Plasmid Preparation:

    Article Title: Monitoring the dynamics of clonal tumour evolution in vivo using secreted luciferases
    Article Snippet: To create a unique EcoRI restriction site, the EcoRI site in the pGIPZ plasmid backbone was destroyed. .. GLuc/CLuc were transferred from pGLuc-Basic (NEB N8082S) and pCLuc-Basic2 (NEB N0317S), respectively, via SbfI/NotI into pPRIME-CMV-dsRed-recipient (Stephen Elledge, Addgene plasmid 11658) generating pPRIME-CMV-GLuc/CLuc. .. In a second step, CMV-GFP in pGIPZ was substituted by CMV-GLuc/CLuc from pPRIME-CMV-GLuc/CLuc via NotI/XbaI digestion and ligation.

    Article Title: HNF1A is a novel oncogene that regulates human pancreatic cancer stem cell properties
    Article Snippet: Biotinylated mouse anti-mouse H-2Kd/H-2Dd clone 34-1-2S was purchased from SouthernBiotech (Birmingham, AL). .. For the Cypridina luciferase construct containing the full-length canonical OCT4 promoter, a 3.9-kbp insert was excised from phOct4-EGFP ( ) by XhoI and BamHI digestion, followed by ligation into pCLuc-Basic2 (New England Biolabs). phOct4-EGFP was a gift from Wei Cui (Addgene plasmid # 38776). .. For the POU5F1/OCT4 LTR construct, a 1.7-kbp insert was amplified from NY5 genomic DNA with the following primers: 5’-ATCTTGGAATTCTGGGCACTCAGTTTATTGTTAGG-3’ and 5’-GGTGGCGGATCCTGTGTTAATCCTCCTCGGGG-3’.

    Article Title: Wnt5a is essential for hippocampal dendritic maintenance and spatial learning and memory in adult mice
    Article Snippet: The fluorescence intensity change is expressed as Δ F / F 0 and the amplitude of fluorescence change (Δ F max / F 0 ) represents the extent of GCaMP3 intensity change. .. To monitor GluN1 promoter activity, the GluN1 – Cypridina reporter construct was generated by subcloning a 1-kb region upstream of the transcription start site of mouse GluN1 into pCLuc Mini-TK 2 Vector (New England Biolabs). .. Neurons were cotranfected with GluN1–Cypridina reporter and a control pSV40-GLuc vector that encodes for Gaussia luciferase under the control of the constitutive SV40 promoter.

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Polymerase Chain Reaction:

    Article Title: Monitoring the dynamics of clonal tumour evolution in vivo using secreted luciferases
    Article Snippet: To obtain lentiviral vectors for inducible expression of luciferases and shRNAs, the pINDUCER10 vector containing PheS stuffer sequence in the EcoRI/XhoI site was modified: tRFP was replaced by GLuc or CLuc from pCR-Blunt II-TOPO-GLuc or pCR-Blunt II-TOPO-CLuc via restriction digest with AgeI/NotI and ligation. .. For pCR-Blunt II-TOPO-GLuc or pCR-Blunt II-TOPO-CLuc, luciferases were amplified by PCR (primer sequences 5′-ACCGGTCAAGCTTGGTACC-3′ and 5′-GCATCTTACTTGGCATGACAGTAAG-3′) from pGLuc-Basic (NEB N8082S) and pCLuc-Basic2 (NEB N0317S). .. For cloning shRNAs, PheS was replaced by shRNAmirs from pGIPZ vectors (Open Biosystems) by EcoRI/XhoI restriction and ligation: shMdm2.1(V2LHS_251529), shMdm2.2(V2LHS_151656), shMdm2.3(V2LHS_379468), shMdm2.4(V2LHS_379469), shPLK1.1(V2LHS_19708), shPLK1.2(V2LHS_19709), shPLK1.3(V2LHS_19711).

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Amplification:

    Article Title: Monitoring the dynamics of clonal tumour evolution in vivo using secreted luciferases
    Article Snippet: To obtain lentiviral vectors for inducible expression of luciferases and shRNAs, the pINDUCER10 vector containing PheS stuffer sequence in the EcoRI/XhoI site was modified: tRFP was replaced by GLuc or CLuc from pCR-Blunt II-TOPO-GLuc or pCR-Blunt II-TOPO-CLuc via restriction digest with AgeI/NotI and ligation. .. For pCR-Blunt II-TOPO-GLuc or pCR-Blunt II-TOPO-CLuc, luciferases were amplified by PCR (primer sequences 5′-ACCGGTCAAGCTTGGTACC-3′ and 5′-GCATCTTACTTGGCATGACAGTAAG-3′) from pGLuc-Basic (NEB N8082S) and pCLuc-Basic2 (NEB N0317S). .. For cloning shRNAs, PheS was replaced by shRNAmirs from pGIPZ vectors (Open Biosystems) by EcoRI/XhoI restriction and ligation: shMdm2.1(V2LHS_251529), shMdm2.2(V2LHS_151656), shMdm2.3(V2LHS_379468), shMdm2.4(V2LHS_379469), shPLK1.1(V2LHS_19708), shPLK1.2(V2LHS_19709), shPLK1.3(V2LHS_19711).

    Construct:

    Article Title: HNF1A is a novel oncogene that regulates human pancreatic cancer stem cell properties
    Article Snippet: Biotinylated mouse anti-mouse H-2Kd/H-2Dd clone 34-1-2S was purchased from SouthernBiotech (Birmingham, AL). .. For the Cypridina luciferase construct containing the full-length canonical OCT4 promoter, a 3.9-kbp insert was excised from phOct4-EGFP ( ) by XhoI and BamHI digestion, followed by ligation into pCLuc-Basic2 (New England Biolabs). phOct4-EGFP was a gift from Wei Cui (Addgene plasmid # 38776). .. For the POU5F1/OCT4 LTR construct, a 1.7-kbp insert was amplified from NY5 genomic DNA with the following primers: 5’-ATCTTGGAATTCTGGGCACTCAGTTTATTGTTAGG-3’ and 5’-GGTGGCGGATCCTGTGTTAATCCTCCTCGGGG-3’.

    Article Title: Wnt5a is essential for hippocampal dendritic maintenance and spatial learning and memory in adult mice
    Article Snippet: The fluorescence intensity change is expressed as Δ F / F 0 and the amplitude of fluorescence change (Δ F max / F 0 ) represents the extent of GCaMP3 intensity change. .. To monitor GluN1 promoter activity, the GluN1 – Cypridina reporter construct was generated by subcloning a 1-kb region upstream of the transcription start site of mouse GluN1 into pCLuc Mini-TK 2 Vector (New England Biolabs). .. Neurons were cotranfected with GluN1–Cypridina reporter and a control pSV40-GLuc vector that encodes for Gaussia luciferase under the control of the constitutive SV40 promoter.

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Ligation:

    Article Title: HNF1A is a novel oncogene that regulates human pancreatic cancer stem cell properties
    Article Snippet: Biotinylated mouse anti-mouse H-2Kd/H-2Dd clone 34-1-2S was purchased from SouthernBiotech (Birmingham, AL). .. For the Cypridina luciferase construct containing the full-length canonical OCT4 promoter, a 3.9-kbp insert was excised from phOct4-EGFP ( ) by XhoI and BamHI digestion, followed by ligation into pCLuc-Basic2 (New England Biolabs). phOct4-EGFP was a gift from Wei Cui (Addgene plasmid # 38776). .. For the POU5F1/OCT4 LTR construct, a 1.7-kbp insert was amplified from NY5 genomic DNA with the following primers: 5’-ATCTTGGAATTCTGGGCACTCAGTTTATTGTTAGG-3’ and 5’-GGTGGCGGATCCTGTGTTAATCCTCCTCGGGG-3’.

    Activity Assay:

    Article Title: Wnt5a is essential for hippocampal dendritic maintenance and spatial learning and memory in adult mice
    Article Snippet: The fluorescence intensity change is expressed as Δ F / F 0 and the amplitude of fluorescence change (Δ F max / F 0 ) represents the extent of GCaMP3 intensity change. .. To monitor GluN1 promoter activity, the GluN1 – Cypridina reporter construct was generated by subcloning a 1-kb region upstream of the transcription start site of mouse GluN1 into pCLuc Mini-TK 2 Vector (New England Biolabs). .. Neurons were cotranfected with GluN1–Cypridina reporter and a control pSV40-GLuc vector that encodes for Gaussia luciferase under the control of the constitutive SV40 promoter.

    Generated:

    Article Title: Wnt5a is essential for hippocampal dendritic maintenance and spatial learning and memory in adult mice
    Article Snippet: The fluorescence intensity change is expressed as Δ F / F 0 and the amplitude of fluorescence change (Δ F max / F 0 ) represents the extent of GCaMP3 intensity change. .. To monitor GluN1 promoter activity, the GluN1 – Cypridina reporter construct was generated by subcloning a 1-kb region upstream of the transcription start site of mouse GluN1 into pCLuc Mini-TK 2 Vector (New England Biolabs). .. Neurons were cotranfected with GluN1–Cypridina reporter and a control pSV40-GLuc vector that encodes for Gaussia luciferase under the control of the constitutive SV40 promoter.

    Subcloning:

    Article Title: Wnt5a is essential for hippocampal dendritic maintenance and spatial learning and memory in adult mice
    Article Snippet: The fluorescence intensity change is expressed as Δ F / F 0 and the amplitude of fluorescence change (Δ F max / F 0 ) represents the extent of GCaMP3 intensity change. .. To monitor GluN1 promoter activity, the GluN1 – Cypridina reporter construct was generated by subcloning a 1-kb region upstream of the transcription start site of mouse GluN1 into pCLuc Mini-TK 2 Vector (New England Biolabs). .. Neurons were cotranfected with GluN1–Cypridina reporter and a control pSV40-GLuc vector that encodes for Gaussia luciferase under the control of the constitutive SV40 promoter.

    Over Expression:

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Knock-Out:

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Mutagenesis:

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    CRISPR:

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Sequencing:

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

    Isolation:

    Article Title: Mutant p53 gain of function underlies high expression levels of colorectal cancer stem cells markers
    Article Snippet: The pRetroSuper-sh-LacZ-puro (sequence: 5′-GTGACCAGCGAATACCTG-3′) was kindly provided by Prof. Reuven Agami. .. The pWZL-blast-mutant p53R175H over-expression vector was constructed as previously described [ ]. pcDNA3-HA-ADH that was used for ALDH1A1 over-expression, was a gift from Steven Johnson (Addgene plasmid # 11610). pX330 p53 vector that was used to knock-out human mutant p53 by CRISPR/Cas9 system (gRNA sequence: 5′-CCATTGTTCAATATCGTCCG-3’) was kindly provided by Dr. Jacob Hanna. pCLuc-Basic2 vector harboring ALDH1A1 promoter was cloned by restriction-free (RF) cloning, in which lkb of ALDH1A1 promoter sequence was isolated from genomic DNA using the following primers: F: 5′-GGATCGGGAGATCTTGGAATTCTGCA-GATAcccatgtaggagttctcttgtg-3′, R: 5′-CCTTAATATGCG AAGGATCCGAGCTCGGagctgctctggccactaaggcc-3′, followed by PCR product clean and secondary PCR using pCLuc-Basic2 vector (NEB) as a template. .. Transfections and retroviral infections Mutant p53 stable knock-down or mutant p53R175H over-expression was performed by stable infections, using ecotropic Phoenix-packaging cells as previously described [ ]. p53 transient knock-down was conducted by cell transfection of specific small interfering RNA (siRNA) oligonucleotides against p53 using Dharmafect3 reagent (Dharmacon). siRNA ON-TARGETplus SMARTpool oligonucleotides were purchased from Dharmacon, including the following sequences: 5′-GAAAUUUGCGUGUGGA- GUA-3′; 5′-GUGCAGCUGUGGGUUGAUU-3′; 5′-GCA-GUCAGAUCCUAGCGUC-3′; 5′-GGAGAAUAUUUCACCCUUC-3′.

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    New England Biolabs pcluc mini tk 2 vector
    Pcluc Mini Tk 2 Vector, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pcluc mini tk 2 vector/product/New England Biolabs
    Average 91 stars, based on 1 article reviews
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    pcluc mini tk 2 vector - by Bioz Stars, 2021-05
    91/100 stars
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