Total RNA was extracted from cells treated with Sal (6 µM), AgNPs (6 µg/mL), or Sal and AgNPs for 24 hours using the Arcturus Picopure RNA Isolation Kit (eBioscience, San Diego, CA, USA), and samples were prepared according to the manufacturer’s instructions. Real-time reverse transcription polymerase chain reaction (RT-PCR) was conducted using a Vill7 (Thermo Fisher Scientific) and SYBR Green as the double-stranded DNA-specific fluorescent dye (Thermo Fisher Scientific). Target gene expression levels were normalized to GAPDH expression, which was unaffected by the treatment. The RT-PCR primer sets are shown in Table 1 . Real-time RT-PCR was performed independently in triplicate for each of the different samples; the data are presented as mean values of gene expression measured in treated sample vs control.
Vill7
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Vill7 is a laboratory instrument designed for performing various analytical tasks. It is a versatile and precise tool that can be used in a wide range of scientific applications. The Vill7 is capable of accurately measuring and analyzing samples, but its specific intended use should not be extrapolated.
Automatically generated - may contain errors
12 protocols using vill7
1
Quantifying Gene Expression Changes
2
Quantifying Gene Expression Changes
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Total RNA was extracted from the cells treated with GO (25 μg/mL), GO-AgNPs (5 μg/mL), or DOX (1 μg/mL) for 24 h using the Arcturus PicoPure RNA isolation kit (Arcturus Bioscience, Mountain View, CA, USA), and then samples were prepared according to the manufacturer’s instructions. Real-time RT-PCR was conducted using a Vill7 (Thermo Fisher Scientific) and SYBR Green as the double-stranded DNA-specific fluorescent dye (Thermo Fisher Scientific). Target gene expression levels were normalized to GAPDH expression, which was unaffected by treatment. The RT-PCR primer sets are shown in Table 1 . Real-time qRT-PCR was performed independently in triplicate for each of the different samples; the data are presented as the mean values of gene expression measured in treated samples versus control.
3
Quantitative Gene Expression Analysis
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Total RNA was extracted from cells treated with rGO-Ag (0.2 µM) or TSA (0.2 µM) or combination of rGO-Ag (0.2 µM) and TSA (0.2 µM) for 24 h using the Arcturus Picopure RNA Isolation Kit (eBioscience, San Diego, CA, USA), and samples were prepared according to the manufacturer’s instructions. Real-time reverse transcription polymerase chain reaction (RT-PCR) was conducted using Vill7 (Thermo Fisher Scientific) and SYBR Green as the double-stranded DNA-specific fluorescent dye (Thermo Fisher Scientific). Target gene expression levels were normalized to GAPDH expression, which was unaffected by the treatment. The RT-PCR primer sets are shown in Table 1 . Real-time RT-PCR was performed independently in triplicate for each of the different samples; the data are presented as mean values of gene expression measured in treated sample vs control.
4
Quantitative Analysis of mRNA Expression
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Extraction and amplification of messenger RNA (mRNA) followed Zhang and Gurunathan.13 (link) According to the manufacturer’s instructions, total RNA was extracted from treated and untreated cells using a Dynabeads mRNA Direct kit (Thermo Fisher Scientific). Real-time quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was conducted using Vill7 (Applied Biosystems, OR, USA) and SYBR green as the double-stranded DNA-specific fluorescent dye (Thermo Fisher Scientific). Target gene-expression levels were normalized to GAPDH gene expression, which was unaffected by Cis, rGO-AgNPs, or Cis plus rGO-AgNP treatment. The real-time qRT-PCR primer sets are shown in Table 1 . Real-time qRT-PCR was performed independently in triplicate for each of the different samples, and data are presented as mean values of gene-expression levels measured in the treated samples versus the controls.
5
Real-Time RT-qPCR Analysis of AgNPs' Effects
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Total RNA was extracted from the cells treated with required concentrations of AgNPs or AgNPs (12.5 μg/mL), or AgNPs (25 μg/mL), or AgNO3 (12.5 μg/mL) for 24 h using the Arcturus PicoPure RNA isolation kit (Arcturus Bioscience, Mountain View, CA, USA), and then samples were prepared according to the manufacturer’s instructions. Real-time RT-qPCR was conducted using a Vill7 (Applied Biosystems, Foster City, CA, USA) and SYBR Green as the double-stranded DNA-specific fluorescent dye (Applied Biosystems). Target gene expression levels were normalized to GAPDH expression, which was unaffected by treatment. The real-time RT-qPCR primer sets are shown in Table 1 . The real-time RT-qPCR was performed independently in triplicate for each of the different samples; the data are presented as the mean values of gene expression measured in treated samples versus the control.
6
Evaluating GEM and AgNPs Effects on Gene Expression
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Cells were treated with 50 nM GEM, 50 nM AgNPs, or a combination of 50 nM GEM and 50 nM AgNPs for 24 h. Total RNA was extracted from the cells using the Arcturus picopure RNA isolation kit (EBioscience, Thermo Fisher Scientific), and samples were prepared according to the manufacturer’s instructions. Real-time reverse transcriptase polymerase chain reaction (RT-PCR) was conducted using Vill7 (Applied Biosystems, Foster City, CA, USA) and SYBR Green as the double-stranded DNA-specific fluorescent dye (Applied Biosystems). The expression levels of target genes were normalized to the expression of GAPDH, which was used as housekeeping gene. The RT-PCR primer sets are shown in Table 1 . Real-time RT-PCR was performed independently in triplicate for each of the different samples; the data are presented as the mean values of gene expression measured in treated samples versus controls.
7
Quantifying AgNP-induced Gene Expression
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Total RNA was extracted from the cells treated with various concentrations of AgNPs for 24 h using the PicoPure RNA isolation kit (Arcturus Bioscience, Mountain View, CA, USA). Samples were prepared according to the manufacturer’s instructions. Real-time RT-qPCR was conducted using a Vill7 (Applied Biosystems, Foster City, CA, USA) and SYBR Green as the double-stranded DNA-specific fluorescent dye (Applied Biosystems). Target gene expression levels were normalized to the expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) expression, which was unaffected by treatment. The real-time qRT-PCR primer sets are shown in Table 2 .
8
Quantification of Gene Expression by qRT-PCR
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According to the manufacturer’s instructions, total RNA was extracted from treated and untreated cells using the Dynabeads mRNA Direct Kit (ThermoFisher, Waltham, MA, USA). Real-time qRT-PCR was conducted using a Vill7 (Applied Biosystems, Foster City, CA, USA) and SYBR Green (Applied Biosystems, Foster City, CA, USA). Target gene expression levels were normalized to the GAPDH gene expression, which was unaffected byAgNPs (1 μM), MS-275 (1 µM), or a combination of AgNPs (1 μM) and MS-275 (1 µM). The real-time qRT-PCR primer sets are shown in Table 1 . Real-time qRT-PCR was performed independently in triplicate, for each of the different samples, and the data are presented as the mean values of the gene expression levels measured in the treated samples versus the controls.
9
Quantifying Gene Expression and mtDNA Levels
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Total RNAs was extracted from blastocysts using Dynabeads mRNA Direct Kit (ThermoFisher, USA) according to the manufacturer's instructions. Real time qRT-PCR was conducted using a Vill7 (Applied Biosystems, OR, USA) and SYBR Green as the double-stranded DNA-specific fluorescent dye (Applied Biosystems, OR, USA). Target gene expression levels were normalized to GAPDH gene expression, which was unaffected by CSNPs treatment. The mitochondrial DNA (mtDNA) content was determined by comparing the ratio of mtDNA to nuclear DNA, as measured by real time qRT-PCR. The real time qRT-PCR primer sets are shown in Tables S2-4 . Real time qRT-PCR was performed independently in triplicate for each of the different samples, and the data are presented as the mean values of the gene expression levels measured in the CSNPs-treated samples versus the controls.
10
Quantitative Analysis of Gene Expression
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Total RNA was extracted from cells, treated with TSA, PdNPs or a combination of both for 24 h, using an Arcturus picopure RNA isolation kit (eBioscience, San Diego, CA, USA); samples were prepared according to the manufacturer’s instructions. Real-time RT-PCR was conducted using a Vill7 (Applied Biosystems, Foster City, CA, USA) and SYBR Green, as the double-stranded DNA-specific fluorescent dye (Applied Biosystems) Target gene expression levels were normalized to GAPDH expression, which was unaffected by treatment. The RT-PCR primer sets are shown in Table 1 . RT-PCR was performed independently in triplicate, for each of the different samples; the data are presented as the mean values of gene expression measured in treated samples versus the control.
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