Light cycler lc480ii

Manufactured by Roche

Sourced in Germany

The Roche LightCycler LC480II is a real-time PCR instrument designed for quantitative and qualitative gene expression analysis. It features a high-performance optical system and a flexible software platform to support a wide range of application needs.

Automatically generated - may contain errors

Lab products found in correlation

7 protocols using light cycler lc480ii

Quantitative Real-Time PCR Analysis of TERRA

Check if the same lab product or an alternative is used in the 5 most similar protocols

TERRA transcript levels were determined from DNA:RNA hybrid RNA samples in the Roche Light Cycler LC480 II (Roche, Mannheim, Germany) using the LightCycler^® 480 SYBR Green I Master Kit (Roche, Cat. No. 04707516001, Mannheim, Germany).
Real-time PCR conditions consisted of an initial denaturation step at 95 °C for 5 min, followed by 40 cycles of 94 °C for 20s, 60 °C for 20s, 72 °C for 45 s and 95 °C for 15 s, and, finally, a melting curve was performed at 67 °C for 1s (melting curve) and cooled at 40 °C for 30 s.
The Ct values of the samples were determined in a Roche Light Cycler LC 480 II Real-Time PCR setup. All samples were run in duplicate to rule out manipulation errors, and Ct values were averaged. β-Actin was used as the house-keeping gene. Ct values obtained at the end of PCR were normalized using the 2^−ΔΔCt method [17 (link),18 (link)].

Kocyigit I., Taheri S., Uysal C., Memis M., Ozayturk S.G., Zararsiz G, & Rassoulzadegan M. (2022). Predicting Progression of Autosomal Dominant Polycystic Kidney Disease by Changes in the Telomeric Epigenome. Cells, 11(20), 3300.

+ Open protocol

+ Expand

Ribo-Zero RNA-Seq Library Preparation

Check if the same lab product or an alternative is used in the 5 most similar protocols

116 ng of total RNA was depleted using the Illumina Ribo-zero rRNA Removal Kit (Bacteria) and purified with Ampure XP beads. Successful depletion was confirmed using Qubit and Agilent 2100 Bioanalyzer. All of the depleted RNA was used as input material for the ScriptSeq v2 RNA-Seq Library Preparation protocol. Following 15 cycles of amplification the libraries were purified using Ampure XP beads. Each library was quantified using Qubit and the size distribution assessed using the Agilent 2100 Bioanalyzer.
The final libraries were pooled in equimolar amounts using the Qubit and Bioanalyzer data. The quantity and quality of each pool was assessed by Bioanalyzer and subsequently by qPCR using the Illumina Library Quantification Kit from Kapa on a Roche Light Cycler LC480II according to manufacturer's instructions. Sequencing was performed at the Centre for Genomic Research, University of Liverpool, on one lane of the Illumina HiSeq 2500 2x125 bp using v4 chemistry (Illumina).

Bronowski C., Mustafa K., Goodhead I., James C.E., Nelson C., Lucaci A., Wigley P., Humphrey T.J., Williams N.J, & Winstanley C. (2017). Campylobacter jejuni transcriptome changes during loss of culturability in water. PLoS ONE, 12(11), e0188936.

+ Open protocol

+ Expand

Illumina RNA Sequencing Protocol

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

The RNA samples were DNase treated and prepared for Illumina sequencing on a HiSeq 2500as described in previous work17 (link). Final libraries were pooled in equimolar amounts using the Qubit and Fragment Analyser data. The quantity and quality of each pool was assessed by the Fragment Analyser and subsequently by qPCR using the Illumina Library Quantification Kit from Kapa on a Roche Light Cycler LC480II according to manufacturer’s instructions. The template DNA was denatured according to the Illumina cBot protocol and loaded at 12 pM concentration. To improve sequencing quality control 1% PhiX was spiked-in. The sequencing was performed on three lanes of an Illumina HiSeq 2500 with version 4 chemistry generating 2 × 125 bp paired end reads.

Bosworth A., Dowall S.D., Garcia-Dorival I., Rickett N.Y., Bruce C.B., Matthews D.A., Fang Y., Aljabr W., Kenny J., Nelson C., Laws T.R., Williamson E.D., Stewart J.P., Carroll M.W., Hewson R, & Hiscox J.A. (2017). A comparison of host gene expression signatures associated with infection in vitro by the Makona and Ecran (Mayinga) variants of Ebola virus. Scientific Reports, 7, 43144.

+ Open protocol

+ Expand

RNA-Seq Protocol for Transcriptome Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total RNA for RNA-Seq was isolated from the wild type strain using mirVana^TM miRNA Isolation Kit (Ambion) according to the protocol for total RNA isolation. Ribosomal RNA was depleted with RiboZero Magnetic kit from Epicentre and, additionally, Ribo Minus^TM Concentrarion Module (Invitrogen). The success of depletion and sample quality was assessed using an Agilent 2100 Bioanalyzer.
RNA-Seq was performed by the Centre for Genomic Research, The University of Liverpool. RNA–Seq libraries were prepared using the Epicentre ScriptSeq v2 RNA-Seq Library Preparation Kit. 17.5 ng of rRNA-depleted RNA, according to quantification by Bioanalyzer, was used as input and following 10 cycles of amplification, libraries were purified using AMPure XP beads. Each library was quantified using a Qubit fluorimeter and the size distribution assessed using the Bioanalyzer. Libraries were pooled in equimolar amounts, based on the Qubit and Bioanalyzer data. The quantity and quality of each pool was assessed by Bioanalyzer and subsequently by qPCR using the Illumina Library Quantification Kit from Kapa on a Roche Light Cycler LC480II. The pool of libraries was sequenced on one lane of the HiSeq 2000 at 2 × 100 bp paired-end sequencing with v3 chemistry.

Chudzicka-Ormaniec P., Macios M., Koper M., Weedall G.D., Caddick M.X., Weglenski P, & Dzikowska A. (2019). The role of the GATA transcription factor AreB in regulation of nitrogen and carbon metabolism in Aspergillus nidulans. FEMS Microbiology Letters, 366(6), fnz066.

+ Open protocol

+ Expand

Next-Gen Sequencing of CRISPR Amplicons

Check if the same lab product or an alternative is used in the 5 most similar protocols

Two separate CRISPR primer (CRISPR1 and CRISPR2 locus) pairs were designed for two first rounds of PCR. Two microliters of template DNA entered a first round of PCR. The primer design incorporates a recognition sequence to allow a secondary nested PCR process. Samples were first purified with Ampure SPRI Beads before entering the second PCR performed to incorporate Illumina adapter sequences. Samples were purified using Ampure SPRI Beads before being quantified using Qubit and assessed for size distribution using the Fragment Analyzer (Agilent). Successfully generated amplicon libraries were taken forward and pooled in equimolar amounts, then size selected with a Pippin Prep machine (Sage Science) using a range of 180–600 bps. The quantity and quality of each pool was assessed by Bioanalyzer and subsequently by qPCR using the Illumina Library Quantification Kit from Kapa on a Roche Light Cycler LC480II according to the manufacturer’s instructions. Template DNA was denatured according to the protocol described in the Illumina cBot User guide (Illumina Document #15006165 v05) and loaded at 12.5 pM concentration. Fragmented PhiX phage genome was added to the sequence library at 15% in order to increase the sequence complexity. The sequencing of each pool was carried out on one lane of an Illumina MiSeq, at 2×250 bp paired-end sequencing with v2 chemistry.

Watson B.N., Capria L., Alseth E.O., Pons B.J., Biswas A., Lenzi L., Buckling A., van Houte S., Westra E.R, & Meaden S. (2024). CRISPR-Cas in Pseudomonas aeruginosa provides transient population-level immunity against high phage exposures. The ISME Journal, 18(1), wrad039.

+ Open protocol

+ Expand

Illumina TruSeq Nano DNA Library Preparation

Check if the same lab product or an alternative is used in the 5 most similar protocols

Libraries were constructed using the TruSeq Nano DNA Sample Preparation Kit (Illumina), and 200 ng of input material. The material was sheared using a Covaris S2 ultrasonicator following the 550 bp insert size protocol. Half (100 ng) of the sheared material was cleaned using 1.6x Sample Purification beads, and half volumes of all reagents were used throughout the protocol. Samples were prepared in a 96-well plate format and size-selected using the Sample Purification beads. Following eight cycles of amplification, the libraries were purified using Sample Purification beads. Each library was quantified using a Qubit (Thermofisher) and the size distribution was assessed using the Agilent 2100 Bioanalyzer. The samples were then pooled and the final library assessed using the Agilent 2100 Bioanalyzer and subsequently subjected to quantitative PCR using the Illumina Library Quantification Kit from Kapa on a Roche Light Cycler LC480II, according to manufacturer's instructions.

Haldenby S., Bronowski C., Nelson C., Kenny J., Martinez-Rodriguez C., Chaudhuri R., Williams N.J., Forbes K., Strachan N.J., Pulman J., Winstanley I.N., Corless C.E., Humphrey T.J., Bolton F.J., O’Brien S.J., Hall N., Hertz-Fowler C, & Winstanley C. (2020). Increasing prevalence of a fluoroquinolone resistance mutation amongst Campylobacter jejuni isolates from four human infectious intestinal disease studies in the United Kingdom. PLoS ONE, 15(1), e0227535.

+ Open protocol

+ Expand

Cricket Head Transcriptome Sequencing

Check if the same lab product or an alternative is used in the 5 most similar protocols

RNA extraction, library preparation and sequencing were performed as described in Pascoal et al. (2016a) . Briefly, we extracted total RNA from cricket heads (n=48; 3 biological replicates for each sex, morph, social treatment and incubator, Table S2) using TRIzol plus RNA purification kits (Life Technologies) and PureLink DNase treatment (Invitrogen), followed by Qubit (Invitrogen) and Bioanalyser (Agilent) quantification and quality control.
We depleted total RNA with RiboZero following the manufacturer's protocol. Purified RNA was checked for depletion and then libraries were constructed using the ScriptSeq protocol (Epicentre). After fragmentation and conversion to cDNA, samples were purified with Ampure XP beads, barcoded, PCR amplified for 14 cycles, and multiplexed. We checked quantity and quality of final pools and performed qPCR using Illumina Library Quantification Kits (Kapa) on a Roche Light Cycler LC480II. Denatured DNA was loaded at 9 pM with 1% fragmented phage PhiX DNA spiked-in, then sequenced on an Illumina HiSeq 2000 (2×100 bp paired end reads).

Pascoal S., Liu X., Fang Y., Paterson S., Ritchie M.G., Rockliffe N., Zuk M, & Bailey N.W. (2018). Increased socially mediated plasticity in gene expression accompanies rapid adaptive evolution. Ecology letters, 21(4).

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!