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THE NEWSLETTER OF ACTIVE MOTIFAPRIL 2014

VOLUME 15 NUMBER 1

Enabling Epigenetics ResearchAntibodies, Kits and Services for Epigenetics & Gene Regulation Research

New to Cancer Epigenetics?
We Can Help.

IN THIS ISSUE
2 New to Cancer Epigenetics?
We Can Help.
4 NEW: Easily Perform ChIP on
Difficult-to-Lyse PBMCs
5 No Sonicator? No Problem!
Perform ChIP Without a Sonicator
6 NEW: Perform Bisulfite Conversion
of DNA in as Little as 1.5 Hours
7 Epigenetic Services: End-to-end
ChIP-Seq and Bisulfite Sequencing
8 NEW: Histone Purification Kits forMore Consistent Sample Prep
9 Recombinant Proteins to Analyze
Histone Modifications
10 NEW: Screen Histone Post-translational Modifications in Multiplex
with our Luminex® Epigenetic Assays
11 Study Post-transcriptional
Regulation Mediated ThroughmiRNA-3´UTR Interactions

2

Over the last decade, epigenetic deregulation has
been increasingly recognized as a hallmark of diseases such as cancer. As a result, many researchers
are studying the roles that epigenetic changes play in
disease. The transition into epigenetics research does
not have to be intimidating because there are an
increasing number of tools available to help
researchers obtain their desired results.
Cancer-linked epigenetic modifications are found genome wide
Epigenetic alterations in cancer
include changes in DNA methylation
and associated histone modifications
that influence the chromatin stateand impact gene expression. The development of methods, such as ChIP
and DNA methylation enrichment
techniques, along with next generation sequencing tools is enabling
researchers to get a better picture of
the genome-wide changes associatedwith cancer and other diseases.
Not every antibody is suitable for
epigenetic techniques
One of the greatest challenges for
epigenetic-based research has been
the lack of antibodies displaying theproper specificity that have been validated for use in techniques such as
ChIP and ChIP-Seq. The problem has
been compounded by numerous antibody suppliers who do not manufacture or test the antibodies they sell,
and who sell them to one anotherand then to researchers. Developing
and manufacturing histone-specificantibodies is challenging because
histones have multiple post-translational modifications along their “tail”
portion involved in gene regulationand disease. Effective antibodies
must clearly differentiate between
these multiple, subtle variances andalso be able to perform in demanding
epigenetic techniques.
What makes a good antibody?
Active Motif has established a validation program for its antibodies toqualify them for their intended use.
This includes ChIP-Seq testing, ChIP
validation as well as a test on thespecificity of our histone antibodies
with a unique MODified™ Histone
Peptide Array. For the full list of ourepigenetics antibodies, please visit
www.activemotif.com/antibodies.

“How do chromatin biology
and epigenetics contribute tocancer development, and what
can be done to treat cancers
caused by epigenetic events?”

www.activemotif.com

Epigenetic Modifications in Cancer

DNA methylation andhydroxymethylation
Aberrant changes in DNA methylation have been well characterized
in a variety of cancers. In general,the cancer epigenome is marked by
global DNA hypomethylation andpromoter-specific DNA hypermethylation, which often leads to silencing
of tumor suppressor genes.

Regulating the histone code
The “histone code” is comprised ofpost-translational modifications that
occur on the histone tails. These
WRITERS
modifications are generated, interpreted and edited by proteins coined acetylases
methylases“Writers”, “Readers” and “Erasers”.
kinases
Aberrant epigenetic regulation canlead to changes in gene expression
and the development of cancer.

The recently characterized DNA
modification 5-hydroxymethylcytosine (5-hmC) has also been linked tocancer. Reductions in both 5-hmClevels and in expression of the TET
enzymes that convert 5-mC to 5-hmC
have been reported in breast, liver,lung, prostate and pancreatic cancers.

The central role of ChIP in
epigenetic studies
ChIP (chromatin immunoprecipitation) is the principal technique used
to map epigenetic marks to individualloci in the genome. By using ChIP, researchers can determine the relationships between histone marks or otherproteins, such as transcription factors,
and gene expression.

Tools for DNA methylation analysis
Due to the roles it plays in development and disease, much researchdepends on the ability to accurately
detect and quantify DNA methylation. Active Motif offers a numberof products specific for this area of
research, including kits and antibodies that enrich for DNA fragmentsthat contain 5-mC and 5-hmC. Manyof our DNA methylation antibodies
have been validated for use in ChIP,
MeDIP and/or immunofluorescence.
For complete details on our DNA
methylation products, please visit us
at www.activemotif.com/dnamt.

sales@activemotif.com

Using the right tools for ChIP
ChIP is a technically challenging
method, and numerous factors cancause it to fail. Therefore, researchers
who are not experts in the techniqueare best served using well-validated
and reliable kits to perform theseassays. Active Motif has developed anumber of kits and accessory reagents tailored to help researcherswith their ChIP experiments. They
provide all the critical components
needed in a single kit along witheasy-to-follow instructions. Thesekits and associated antibodies havebeen used in hundreds of labs andcited in thousands of papers in peer
reviewed journals. For a complete listof kits and reagents for ChIP, please
visit us at www.activemotif.com/chip.

READERS
bromodomain
chromodomain
PHD finger
WD40 repeat+

ERASERS
deacetylases
demethylases
phosphatases

Let us do the work for you:
Active Motif Epigenetic Services
Reproducibly generating high-quality,interpretable data from ChIP experiments can be challenging as it
requires prior knowledge of working
antibodies, optimized protocols forvarious cell types and knowledge ofcell type-specific binding sites. Addin the technical and bioinformatics
challenges associated with generating whole-genome data sets, and
ChIP-Seq may literally be beyond
your reach. That is why our Epigenetic
Services team provides a wide varietyof ChIP services, making it possible
for you to utilize our expertise andresearch tools without having to be
an expert in the techniques yourself.
To find out more, or to get a quote,go to www.activemotif.com/services.

“Epigenetic applications require the use of antibodies withthe proper specificity that have been validated foruse in techniques such as ChIP, ChIP-Seq and MeDIP.”

3

Chromatin Immunoprecipitation

NEW

Easily Perform ChIP on Difficult-to-Lyse PBMCs
Active Motif’s new ChIP-IT® PBMC Kit overcomes the challenges ofgenerating quality chromatin for use in ChIP from peripheral bloodmononuclear cells (PBMCs), such as lymphocytes (T and B cells) andmonocytes, that are highly resistant to lysis using conditions normallysuitable for other cells. The ChIP-IT PBMC Kit offers a complete
solution for ChIP by providing optimized buffers and lysis conditions
together with our highly sensitive ChIP reagents, ensuring successful
ChIP reactions even when working with problematic cells like PBMCs.

What’s in the box?
The ChIP-IT PBMC Kit contains
optimized buffers and protocols that
improve the efficiency of PBMC cell
lysis, enabling extraction of quality
chromatin for ChIP. It also contains
our high sensitivity ChIP reagents for
performing 16 chromatin immunoprecipitation reactions using our low
background Protein G agarose beads
and ChIP filtration columns for quick,easy capture and washing steps.

4

qPCR analysis simplified
For downstream analysis of ChIP-IT
PBMC generated data, we highly recommend our ChIP-IT qPCR Analysis
Kit. It includes control reagents andspreadsheets designed to simplify

the analysis and interpretation ofqPCR data, enabling you to assess
the quality of your ChIP’d DNA prior
to investing in expensive sequencing
experiments. For more information,
visit www.activemotif.com/pbmc.

PBMC ChIP on CD8+ T Cells-''
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PBMCs, including lymphocytes(T cells, B cells and NK cells) andmonocytes, constitute a criticalcomponent of the peripheral immune
system. Because of the importance of
PBMCs in immunology and infectious
disease research, a robust method to
extract quality chromatin from thesecells for ChIP and ChIP-Seq analysis
is highly warranted. Active Motif applied its years of experience working
with these difficult-to-lyse sample
types into developing an optimized
chromatin preparation method thatyields high-quality chromatin from
PBMCs. Used with our highly sensitive
ChIP reagents, this new method will
improve your results and help you
achieve better ChIP data.

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Figure 1: ChIP-IT PBMC shows H3K9ac enrichment at the ACTB and GAPDH promoters.
The ChIP-IT PBMC Kit was used to prepare chromatin from CD8+ T cells and perform ChIP
using a Histone H3K9ac antibody. Following enrichment, qPCR was performed using the
ChIP-IT qPCR Analysis Kit to normalize the data. Enrichment of H3K9ac was observed with
the ACTB-2 (Catalog No. 71005) and GAPDH-2 (Catalog No. 71006) Positive Control Primer Sets,
but the Negative Control Primer Set (Catalog No. 71001) showed no enrichment.

Product

FormatCatalog No.

ChIP-IT® PBMC

16 rxns

53042

ChIP-IT® qPCR Analysis Kit

10 rxns

53029

www.activemotif.com

Chromatin Preparation

No Sonicator? No Problem!
Perform ChIP Without a Sonicator
Active Motif’s ChIP-IT® Express Enzymatic Kit makes it
possible to perform chromatin immunoprecipitation (ChIP)
experiments quickly and easily from cultured cell samples
without the need for a sonicator. It uses a proprietary
enzymatic shearing cocktail to reproducibly shear DNA
into 200-1000 bp fragments that are ideal for ChIP.
Why another ChIP Kit?
Although sonication is an effective
method to shear DNA, not everybody
owns a sonicator. For new users, it
can be difficult to optimize sonication parameters and get reproducible
results due to complications such as
emulsification and overheating.

Because of this, Active Motif has developed a robust and extremely userfriendly method to shear chromatin
for ChIP using enzymatic digestion.
The ChIP-IT Express Enzymatic Kit
uses a proprietary Enzymatic Shearing
Cocktail that quickly shears DNA into
200-1000 bp fragments (Figure 1).
Because enzymatic shearing is solely
time and temperature dependent, theproblems associated with sonication
are eliminated. Enzymatic shearing isa good choice for users who do notplan to do enough ChIP to make itworthwhile to invest the time andmoney required to purchase and learn
how to use a sonicator.
What’s in the box?
The ChIP-IT Express Enzymatic Kits
include Protein G Magnetic Beads, an
enzymatic shearing cocktail, proteaseinhibitors, lysis enzymes and buffers.

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The kit provides sufficient reagentsfor performing 25 ChIP reactions.
Shearing components are included
to optimize shearing conditions andthen make 5 preparations of shearedchromatin from three 15 cm plates(4.5 x 107 cells); each preparation
yields enough chromatin to perform
approximately 6 ChIP reactions.

ChIP companion products help
make chromatin IP even easier
Active Motif offers a number of ChIP
Accessory Products that are designed
to make it even easier for you to perform successful ChIP when working
with our ChIP-IT Kits. These include
ChIP qPCR primer sets, ChIP-validated
antibodies, ChIP-IT Control Kits andChIP DNA cleanup kits.

For more information, please visit us
at www.activemotif.com/chip.

1000 bp
500 bp100 bp
1

2

3

4

5

Figure 1: Gel analysis of sheared chromatin
prepared using the ChIP-IT Express Enzymatic Kit.
Lane 1: 100 to 1000 bp ladder.
Lane 2: Unsheared HeLa DNA.
Lanes 3-5: HeLa DNA incubated with Enzymatic
Shearing Cocktail for 5, 10 and 15 minutes.

ChIP Express Enzymatic advantages• No sonication equipment
required• User-friendly protocol• Simplifies optimization offragment size
• Mild enzymatic treatment
preserves chromatin andepitope integrity• Perform ChIP in just 1 day

Product

FormatCatalog No.

ChIP-IT® Express Enzymatic

25 rxns

53009

ChIP-IT® Express Enzymatic Shearing Kit

10 rxns

53035

Chromatin IP DNA Purification Kit

50 rxns

58002

ChIP-IT® Control qPCR Kit – Human

5 rxns53026

5

Bisulfite Conversion

NEW

Perform Bisulfite Conversion of DNA in as Little as 1.5 Hours
Active Motif’s new Bisulfite Conversion Kit simplifies the bisulfite treatment
of DNA while minimizing DNA fragmentation, degradation and sample loss
during the treatment and cleanup steps. A thermal cycler is used to denature the DNA and perform bisulfite conversion in as little as 1.5 hours. The
included spin columns combine desulfonation and DNA purification into asingle step. The purified, converted DNA is ideal for PCR amplification andsequencing, endonuclease digestions and other downstream applications.

Highlights• Rapid conversion of DNA in as
little as 1.5 hours• Minimal DNA fragmentation,
degradation and loss
• > 99% conversion efficiency• Easily convert 200 pg – 2 µg DNA• Conversion-specific PCR primers
included to validate results
How does it work?
Bisulfite conversion and subsequent
DNA sequencing is considered thegold standard in DNA methylation
analysis as it provides single base-pairresolution of the DNA methylation
profile. The conversion reaction occurs as a three-step deamination ofcytosine residues into uracil. As only
unmethylated cytosine residues are
susceptible to bisulfite conversion,
the original methylation state of the
DNA can be determined (Figure 1).
What’s in the box?
The Bisulfite Conversion Kit contains
proteinase K for pre-treatment of
the DNA, optimized conversion andpurification reagents, an easy-to-use
protocol and conversion-specific PCRprimers suitable for use with human6

Bisulfite Treatment Converts Unmethylated Cytosines into Uracils
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Figure 1: Bisulfite treatment converts unmethylated cytosine residues into uracils, which are then converted tothymines following PCR amplification. Methylated cytosines are protected, so they remain unchanged.

Step-by-step Bisulfite Conversion=[dec_Y:D7
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and mouse samples. Because the conversion-specific primer pair produces
a PCR product only if conversion has
occurred, the primers can be used tovalidate results before starting sequencing or other analysis methods.
For more information, please visit
www.activemotif.com/bis-conv.

Product

FormatCatalog No.

Bisulfite Conversion Kit

50 rxns

55016

www.activemotif.com

Contract Research Services

Epigenetic Services: End-to-end ChIP-Seq andBisulfite Sequencing Assays
Active Motif is committed to being the leader in the field of epigenetics andto making whole-genome epigenetics and transcriptional regulation data
accessible to everyone. That is why, in addition to our wide range of kits andantibodies for chromatin, DNA methylation and transcription research, our
Epigenetic Services team performs end-to-end services for ChIP, ChIP-Seq,bisulfite sequencing and more. Simply send us your samples and we’ll
provide you with high-quality data that will propel your research forward.
ChIP antibody validation:
The #1 challenge in ChIP-Seq
Researchers performing ChIP-Seq
encounter many challenges in thismultistep protocol. However, finding
a good antibody may be the biggest
hurdle because so few function in
ChIP-Seq. Therefore, Active Motif
offers ChIP validation of any custom
or commercially available antibody.
The quickest, most reliable
path to antibody validation' :ljkfd\ij\e[j]`o\[fi]ifq\e
Z\ccfik`jjl\jXdgc\j%( 8Zk`m\Dfk`]gi\gXi\jZ_ifdXk`e%) 8Zk`m\Dfk`]g\i]fidj:_@G%* :_@G$J\hc`YiXi`\jXi\ZfejkilZk\[%
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performing ChIP
• 25% of all transcription factor
antibodies• 80% for histone modification
antibodies

Targeted Next-Gen Bisulfite
Sequencing: DNA methylation atbase pair resolution
For researchers that have alreadyidentified differentially methylated
regions or key genes of interest using
a genome-wide methylation analysis
approach, the next step is validationof those regions.

Active Motif’s Targeted Next-Gen
Bisulfite Sequencing Service offersa high-throughput solution. This
assay combines multiple bisulfiteconverted regions from several dif
different samples into a single Next-Gen
sequencing reaction, resulting in morestatistically significant data at a lower
cost than traditional Sanger-based
bisulfite sequencing.
Targeted Next-Gen Bisulfite
Sequencing Service includes• Bisulfite conversion• Primer design and testing• PCR amplification• Barcoded library generation• DNA sequencing• Data analysis

For more information, or to request a quote, please go to www.activemotif.com/services.

sales@activemotif.com

7

Histone Purification

NEW

Histone Purification Kits for More Consistent Sample Prep
Improve the consistency and reliability of your analysis of histone
modifications by using purified histone samples. Purification ofhistones eliminates contaminating acid-insoluble cellular proteinsthat are left behind with standard acid extraction protocols.
Removal of these impurities also prevents additional alterations
to histone modifications, ensuring you have the highest quality
histone samples for use in your downstream analysis.

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in various downstream applications
such as Western blot, mass spectrometry and in high-throughput methods,
including our Histone Modification
ELISAs (Figure 1) and Histone H3 PTM
Multiplex Assay (page 10).

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The pure advantage
Active Motif is the only company tooffer a method for isolating purified
core histones from cell and tissue
samples that preserves theirpost-translational modifications.

The proprietary buffers and spin columns included in our purification kits
are available in multiple formats, so
you can choose the one that best fits
the needs of your experiment (Table 1).
Isolated histones are suitable for use

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Advantages• Exclusivity – only commercial kitsavailable for histone purification• Quality – enhanced histone
purity and preservation ofhistone modifications• Scalability – 3 throughput formats
available to suit your needs• Flexibility – works with cell or
tissue samples

Figure 1: Active Motif’s histone
purification method improves histoneyield over crude acid extraction.
The Total Histone H3 ELISA Kit(Cat. No. 53110) was used to generate a standard curve with the kit’s
Recombinant Histone H3 protein (red).
The kit was then used to quantify
the amount of Total H3 in purified
histone samples obtained using theHistone Purification Mini Kit and theHistone Purification Microplate Kit ascompared to samples isolated using
crude acid extraction.

Histone Purification Kit

Histone Purification Mini Kit

Histone Purification Microplate Kit

40025

40026

40027

Application• Low
throughput• Highsampleamounts

FractionsSingle fraction, or separated into
H2A/H2B & H3/H4 fractions

Single fraction containing
H2A, H2B, H3 & H4Single fraction containing
H2A, H2B, H3 & H4

10

20

96

# of purifications• Mediumthroughput• Mid-range
sampleamounts• Highthroughput• Low
sampleamounts

Table 1: Specifications of Active Motif’s various Histone Purification Kits. For more information, please got to www.activemotif.com/histonepur.

8

www.activemotif.com

Recombinant Proteins

Recombinant Proteins to Analyze Histone Modifications
Active Motif offers a comprehensive selection of recombinant histone
modifying enzymes, as well as unmodified and modified histones, forepigenetic drug discovery applications. Our patented synthesis technologies
enable Active Motif to be the exclusive source for many full-length modified
histones. We also offer a broad selection of bromodomains, HATs, HDACs,
HMTs and HDMs for use in studies of enzyme kinetics, inhibitor screens,selectivity profiling and structural analysis.

Assay-ready proteins include:• The largest selection of Modified
and Unmodified Histones• The largest selection of
Bromodomain proteins• Acetyltransferases & Deacetylases• Methyltransferases & Demethylases
Assay-ready recombinant proteinsIf you are analyzing histone modifications or Readers, Writers and Erasers
as part of your research, there is noneed to waste your valuable time andresources producing protein substrates and enzymes for use in yourassays. Active Motif offers a comprehensive selection of high-quality
recombinant proteins, enzymes anddomains for analysis of histone
modifications that can readily beincorporated into your assays to help
accelerate your research.
Proteins for Biochemical Assays• Enzyme activity assays• Inhibitor screens• Selectivity profiling• Structural analysis• TR-FRET

sales@activemotif.com

With over 300 purified assay-readyrecombinant proteins and a large
selection of recombinant histones
and histone modifying enzymes – including bromodomains, HATs, HDACs,
HMTs and HDMs – you are sure tofind what you need to develop more
efficient assays for your drug discovery and development programs.
For the complete list of purified
recombinant proteins available, please
go to www.activemotif.com/proteins.

modifications in a native chromatin
environment. By using the most biologically relevant substrate, you canensure the most accurate result.

Figure 1: Which substrate is right for your assay?
Choose the correct substrate to achieve the best
results. Opinion leaders in epigenetics recognize thepower of reconstituting recombinant chromatin for
creating biologically relevant substrates.

Full-length modified histones
Your biochemical assay can betterselect and validate activity when thesystem mimics actual cellular biology(Figure 1). To help you make thecorrect choice of histone substrate,
Active Motif offers the largest
selection of recombinant histones
with over 60 different H3 and H4
histones to choose from.

Exclusive, comprehensive portfolio
Because of our patented Expressed
Protein Ligation (EPL) and Methylated
Lysine Analog (MLA) synthesis
technologies, Active Motif offers themost exclusive and comprehensive
set of full-length modified histones.

Our recombinant histones can beused as stand-alone substrates or
combined with recombinant H2A andH2B to generate nucleosomes. Oligonucleosomes can be generated using
our Chromatin Assembly Kit (Cat. No.
53500). It is important to analyze theeffects of DNA sequence and histone

Our exclusive collection of modified
recombinant histones offers a variety
of site- and degree-specific methyl,acetyl and phosphoryl modifications,as well as biotinylated histones tochoose from. For a complete list ofavailable histones, please visit us atwww.activemotif.com/recombhis.

9

Histone H3 PTM Multiplex Assays

NEW

Screen Histone Post-translational Modifications in
Multiplex with our Luminex® Epigenetic Assays
The Histone H3 PTM Multiplex Assay is the first commercial
multiplex assay for screening global changes in the levels of histone post-translational modifications (PTMs). The assay enables
simultaneous analysis of multiple modifications in a single wellusing smaller sample amounts, in less time and at a fraction of
the cost of traditional methods, such as Western blots.

Each bead emits a specific, uniquefluorescent signal, enabling the multiplexing of multiple analytes in thesame sample. You can also measure
the magnitude of the streptavidinphycoerythrin signal to provide areal-time readout of
PTM levels (Figure 2).

The Histone H3 PTM Multiplex Assayworks as a solution-based suspension
sandwich “ELISA” to analyze relative
levels of histone H3 modifications.
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A complete assay
requires purchase ofboth the Histone H3
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containing buffers andreagents required toperform the assay as

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How does it work?
Active Motif partnered with Luminex,
the industry leader in multiplexing, todevelop the first multiplex epigenetic
assay for high-throughput analysis ofhistone modifications.

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Histones are captured using fluorescent labeled magnetic beads that are
conjugated to antibodies recognizing specific PTMs in histone tails. A
biotinylated H3 C-terminal antibody
is used to capture histones. Streptavidin-phycoerythrin is then added
to bind the biotinylated antibody toprovide a signal of the binding events(Figure 1). Inclusion of the Histone H3
Total beads in a multiplex enables
normalization of values against total
histone H3 levels so you can determine the relative amounts of eachhistone modification in each sample.

Advantages• Multiplex analysis of up to 13 PTMs
• Requires only nanograms of sample• 3 hour assay• Ability to normalize & compare
relative PTM levels across samples• Higher throughput, faster andless expensive than Western
blotting or AlphaLISA

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Figure 2: Single-well, multiplexed data from theHistone H3 PTM Assay simultaneously reveals specific,
off-target and non-specific effects of VPA inhibitor.

well as the Ab-conjugated bead(s)for your analytes of interest. To
learn more, please visit us atwww.activemotif.com/luminex.

www.activemotif.com

m i c r o R N A s & 3´ U n t r a n s l a t e d R e g i o n s

Study Post-transcriptional Regulation
Mediated Through miRNA-3´UTR Interactions
MicroRNAs (miRNAs) are small, non-coding RNA molecules thatplay a large role as post-transcriptional regulators. miRNAs
repress expression by targeting the 3´UTR (untranslated) regionof mRNA transcripts. The interaction of a miRNA with a 3´UTRresults in either inhibition of translation or increased
degradation of the targeted transcript.

Applications• Understand the mechanism
by which a gene is induced or
repressed• Measure the impact of a3´UTR on post-transcriptional
gene regulation• Validate the targets of amiRNA or siRNA• Measure the effects ofsequence variants on
3´UTR or miRNA function
Complete regulation solution
The LightSwitch™ Luciferase Assay
System is ideal for performing miRNAtarget validation and assessing thefunctional impact of miRNA-3´UTRinteractions. It includes the 3´UTR
Reporter GoClone™ Collection ofover 12,000 human 3´UTRs cloned
from the human genome into readyto-transfect LightSwitch reporter
vectors. Combined with our large
collections of miRNA Mimics & Inhibitors, you have everything needed to
study miRNA-3´UTR interactions,validate miRNA targets, measure RNA
stability, translation efficiency andthe functional impact of miRNAs on agene-by-gene basis (Figure 1).

sales@activemotif.com

Optimized luciferase gene,substrate and assay reagentsThe LightSwitch System was designed to ensure you get the best
results possible. LightSwitch vectors
utilize RenSP, an optimized Renillaluciferase gene that was engineered
for maximum brightness and minimal
background. In addition, LightSwitch
Assay Kits feature a novel, proprietary
assay substrate and an optimized lysis
buffer that provide high sensitivity
over a broad dynamic range.
Get your clones of interest fast
To get started, use our convenientonline search tool to find yourelements of interest. As GoClone
constructs are supplied transfectionready, you can begin your experiments immediately without theneed to clone or prepare DNA.
Accelerate your research
Alternatively, our Custom Services
team can quickly and economicallyclone any fragment from the human, mouse or rat genome into any
LightSwitch vector for you. We alsooffer miRNA Target Validation and
Sequence Variant Assay services. For
more information, please visit us atwww.activemotif.com/lightswitch.

Human genome
Gene of interest
RenSP
Luciferase

3´UTR
Pre-cloned intoa Reporter
VectorLightSwitch™
3´UTR
ReportermiRNA Mimic
RISC
complex

1. Co-transfectvector with
miRNA mimics
or inhibitors.

Hybrid mRNA

Light signal

2. Measure luciferase activity to determine
the effect on translation of miRNA binding
to the hybrid luciferase-3´UTR mRNA.
Figure 1: LightSwitch 3´UTR Reporter Assays.
A 3´UTR cloned into a LightSwitch 3´UTR Reporter
vector is co-transfected with miRNA mimics orinhibitors. The relative amounts of light produced incells with and without miRNA are used to measure
the effect of the miRNA-3´UTR interaction on translation of the hybrid RenSP-3´UTR mRNA transcript.

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Enabling Epigenetics Research

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