1.普通质粒表达载体:pTRE-F.Luc
载体特征 1) 在转录调控因子结合位点(TRE)上游插入了人造poly(A)终止信号,可以有效的降低背景; 2) 利用萤火虫荧光素酶基因(Fluc)可对信号活性做出准确的定量分析; 3) 利用Hygromycin抗性基因筛选稳定表达的细胞系。
2.
1) 反向克隆TRE-pA-Fluc.copGFP表达框,可有效减少因LTR启动子造成的背景;
2) 利用萤火虫荧光素酶基因(Fluc)可对信号活性做出准确的定量分析;
3) 通过copGFP绿色荧光标记蛋白可实现对信号通路活性的快速观测;
4) 可以直接转染,也可包装成病毒后感染;
5) 利用Puromycin抗性基因筛选稳定表达的细胞系。
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Pathway reporter实例
1. NFAT-FLuc
转染后24h用10ng/mL PMA和1µM ionomycin处理细胞,用Luciferase Assay进行检测。
2. NFkB-Fluc2
转染后24h用50ng/mL TNF-α处理细胞,用Luciferase Assay进行检测。
3. pTGFβ-Fluc2
转染后24h用25ng/µL TGFβ处理细胞,用Luciferase Assay进行检测。
4. pAP-1-Fluc2
转染后24h用10ng/mL PMA处理细胞,用Luciferase Assay进行检测。
5. HIF1a-Fluc2
转染后24h用250µM CoCl2处理细胞8h,用Luciferase Assay进行检测。
Pathway reporter载体
1. pTRE-Fluc2系列:
已完成45个pathway reporter,详情见下表:
Vector Name |
Pathway |
Transcription Factor(TF) |
pAhR-Fluc2 |
Xenobiotic |
Xenobiotic Response Element |
pAR-Fluc2 |
Androgen |
Androgen Receptor |
pAP-1-Fluc2 |
MAPK/JNK |
AP-1 |
pATF4/3/2-Fluc2 |
Amino Acid Deprivation |
ATF4/3/2 |
pATF6-Fluc2 |
ATF6 |
ATF6 |
pC/EBP-Fluc2 |
C/EBP |
C/EBP |
pCBF/NF-Y/YY1-Fluc2 |
ER Stress |
CBF/NF-Y/YY1 |
pCREB-Fluc2 |
cAMP/PKA |
CREB |
pE2F/DP1-Fluc2 |
Cell Cycle |
E2F/DP1 |
pEGR1-Fluc2 |
EGR1 |
EGR1 |
pElk-1/SRF-Fluc2 |
MAPK/ERK |
Elk-1/SRF |
pER-Fluc2 |
Estrogen Receptor |
Estrogen Receptor (ER) |
pFOXO-Fluc2 |
PI3K/AKT |
FOXO |
pGATA-Fluc2 |
GATA |
GATA |
pGLI-Fluc2 |
Hedgehog |
GLI |
pGR-Fluc2 |
Glucocorticoid Receptor |
Glucocorticoid Receptor (GR) |
pHNF4-Fluc2 |
Hepatocyte Nuclear Factor 4 |
HNF4 |
pHSF-Fluc2 |
Heat Shock Response |
HSF |
pHIF-1-Fluc2 |
Hypoxia |
Hypoxia-inducible factor-1 (HIF-1) |
pIRF1-Fluc2 |
Interferon Regulation |
IRF1 |
pKLF4-Fluc2 |
KLF4 |
KLF4 |
pLXRa-Fluc2 |
Liver X Receptor |
LXRa |
pMEF2-Fluc2 |
MEF2 |
MEF2 |
pMTF1-Fluc2 |
Heavy Metal Stress |
MTF1 |
pMyc/Max-Fluc2 |
c-myc |
Myc/Max |
pNanog-Fluc2 |
Nanog |
Nanog |
pNFAT-Fluc2 |
PKC/Ca++ |
NFAT |
pNFκB-Fluc2 |
NFκB |
NFκB |
pNrf2 & Nrf1-Fluc2 |
Antioxidant Response |
Nrf2 & Nrf1 |
pOct4-Fluc2 |
Oct4 |
Oct4 |
pp53-Fluc2 |
p53/DNA Damage |
p53 |
pPax6-Fluc2 |
Pax6 |
Pax6 |
pPPAR-Fluc2 |
PPAR |
PPAR |
pProgesterone Receptor-Fluc2 |
Progesterone |
Progesterone Receptor |
pRBP-Jk-Fluc2 |
Notch |
RBP-Jk |
pRAR-Fluc2 |
Retinoic Acid Receptor |
Retinoic Acid Receptor (RAR) |
pRetinoid X Receptor-Fluc2 |
Retinoid X Receptor |
Retinoid X Receptor |
pSMAD2/SMAD3/SMAD4-Fluc2 |
TGFβ |
SMAD2/SMAD3/SMAD4 |
pSox2-Fluc2 |
Sox2 |
Sox2 |
pSP1-Fluc2 |
SP1 |
SP1 |
pSTAT1/STAT1-Fluc2 |
Interferon Gamma |
STAT1/STAT1 |
pSTAT1/STAT2-Fluc2 |
Type I Interferon |
STAT1/STAT2 |
pSTAT3-Fluc2 |
STAT3 |
STAT3 |
pTCF/LEF-Fluc2 |
Wnt |
TCF/LEF |
pVitamin D Receptor-Fluc2 |
VDRE |
Vitamin D Receptor |
pCMV-Fluc2-Negative Control |
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pFluc2-Positive Control |
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注明:如需包装慢病毒报告载体,请联系本公司销售部。
2. pL-TRE-Luc.copGFP系列:
已完成的pathway reporter见下表:
Vector Name |
Pathway |
Transcription Factor(TF) |
pL-AhR-Fluc2 |
Xenobiotic |
Xenobiotic Response Element |
pL-AR-Fluc2 |
Androgen |
Androgen Receptor |
pL-AP-1-Fluc2 |
MAPK/JNK |
AP-1 |
pL-ATF4/3/2-Fluc2 |
Amino Acid Deprivation |
ATF4/3/2 |
pL-ATF6-Fluc2 |
ATF6 |
ATF6 |
pL-C/EBP-Fluc2 |
C/EBP |
C/EBP |
pL-CBF/NF-Y/YY1-Fluc2 |
ER Stress |
CBF/NF-Y/YY1 |
pL-CREB-Fluc2 |
cAMP/PKA |
CREB |
pL-E2F/DP1-Fluc2 |
Cell Cycle |
E2F/DP1 |
pL-EGR1-Fluc2 |
EGR1 |
EGR1 |
pL-Elk-1/SRF-Fluc2 |
MAPK/ERK |
Elk-1/SRF |
pL-ER-Fluc2 |
Estrogen Receptor |
Estrogen Receptor (ER) |
pL-FOXO-Fluc2 |
PI3K/AKT |
FOXO |
pL-GATA-Fluc2 |
GATA |
GATA |
pL-GLI-Fluc2 |
Hedgehog |
GLI |
pL-GR-Fluc2 |
Glucocorticoid Receptor |
Glucocorticoid Receptor (GR) |
pL-HNF4-Fluc2 |
Hepatocyte Nuclear Factor 4 |
HNF4 |
pL-HSF-Fluc2 |
Heat Shock Response |
HSF |
pL-HIF-1-Fluc2 |
Hypoxia |
Hypoxia-inducible factor-1 (HIF-1) |
pL-IRF1-Fluc2 |
Interferon Regulation |
IRF1 |
pL-KLF4-Fluc2 |
KLF4 |
KLF4 |
pL-LXRa-Fluc2 |
Liver X Receptor |
LXRa |
pL-MEF2-Fluc2 |
MEF2 |
MEF2 |
pL-MTF1-Fluc2 |
Heavy Metal Stress |
MTF1 |
pL-Myc/Max-Fluc2 |
c-myc |
Myc/Max |
pL-Nanog-Fluc2 |
Nanog |
Nanog |
pL-NFAT-Fluc2 |
PKC/Ca++ |
NFAT |
pL-NFκB-Fluc2 |
NFκB |
NFκB |
pL-Nrf2 & Nrf1-Fluc2 |
Antioxidant Response |
Nrf2 & Nrf1 |
pL-Oct4-Fluc2 |
Oct4 |
Oct4 |
pL-p53-Fluc2 |
p53/DNA Damage |
p53 |
pL-Pax6-Fluc2 |
Pax6 |
Pax6 |
pPPAR-Fluc2 |
PPAR |
PPAR |
pL-Progesterone Receptor-Fluc2 |
Progesterone |
Progesterone Receptor |
pL-RBP-Jk-Fluc2 |
Notch |
RBP-Jk |
pL-RAR-Fluc2 |
Retinoic Acid Receptor |
Retinoic Acid Receptor (RAR) |
pL-Retinoid X Receptor-Fluc2 |
Retinoid X Receptor |
Retinoid X Receptor |
pL-SMAD2/SMAD3/SMAD4-Fluc2 |
TGFβ |
SMAD2/SMAD3/SMAD4 |
pL-Sox2-Fluc2 |
Sox2 |
Sox2 |
pL-SP1-Fluc2 |
SP1 |
SP1 |
pL-STAT1/STAT1-Fluc2 |
Interferon Gamma |
STAT1/STAT1 |
pL-STAT1/STAT2-Fluc2 |
Type I Interferon |
STAT1/STAT2 |
pL-STAT3-Fluc2 |
STAT3 |
STAT3 |
pL-TCF/LEF-Fluc2 |
Wnt |
TCF/LEF |
pL-Vitamin D Receptor-Fluc2 |
VDRE |
Vitamin D Receptor |
pL-Fluc2-Negative Control |
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pL-Fluc2-Positive Control |
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