INSTRUCTION MANUAL
HiScribe™ T7 Quick High Yield RNA Synthesis Kit
NEB #E2050S/L

50/250 reactions
Version 6.1_9/24

Table of Contents
Introduction ..................................................................................................................................................................................................2
DNA Template Preparation ........................................................................................................................................................................2
Plasmid Templates ................................................................................................................................................................................2
PCR Templates .....................................................................................................................................................................................3
Synthetic DNA Oligonucleotides .........................................................................................................................................................3
RNA Synthesis Protocols ............................................................................................................................................................................3
Standard RNA Synthesis .......................................................................................................................................................................3
Capped RNA Synthesis .........................................................................................................................................................................4
RNA Synthesis with Modified Nucleotides ..........................................................................................................................................5
Purification of Synthesized RNA ................................................................................................................................................................6
LiCl Precipitation ...................................................................................................................................................................................6
Phenol:Chloroform Extraction and Ethanol Precipitation ....................................................................................................................6
Spin Column Chromatography .............................................................................................................................................................6
Gel Purification .....................................................................................................................................................................................7
Evaluation of Reaction Products .................................................................................................................................................................7
Quantification by UV Light Absorbance ..............................................................................................................................................7
Analysis of Transcription Products by Gel Electrophoresis .................................................................................................................7
Troubleshooting ..........................................................................................................................................................................................8
Control Reaction ...................................................................................................................................................................................8
Low Yield of Full-length RNA .............................................................................................................................................................8
Addition of DTT ....................................................................................................................................................................................8
Low Yield of Short Transcript ..............................................................................................................................................................8
RNA Transcript Smearing on Denaturing Gel ......................................................................................................................................8
RNA Transcript of Larger Size than Expected .....................................................................................................................................8
RNA Transcript of Smaller Size than Expected ...................................................................................................................................8
Ordering Information ..................................................................................................................................................................................9Revision History ........................................................................................................................................................................................10

The HiScribe T7 Quick High Yield RNA Synthesis Kit Includes:
All kit components should be stored at –20°C. Each kit contains sufficient reagents for 50 x 20 µl reactions (S size) or
250 x 20 µl reactions (L size). Each standard reaction yields up to 180 µg of unmodified RNA from 1 µg control template.
NTP Buffer Mix (20 mM each NTP)
T7 RNA Polymerase Mix
FLuc Control Template (0.5 µg/µl)
DNase I (2 units/µl)
LiCl Solution (7.5 M LiCl, 10 mM EDTA)
Dithiothreitol (DTT) (0.1 M)

1Required Materials Not Included:
DNA Template
Thermocycler or 37°C incubator
Nuclease-Free Water
Buffer- or water-saturated phenol:chloroform
Ethanol
3 M Sodium Acetate, pH 5.2
5 M Ammonium Acetate
Spin Columns (See Monarch® RNA Cleanup Kits, NEB #T2030, #T2040, #T2050)
Gels, running buffers and gel box

Introduction
The HiScribe T7 Quick High Yield RNA Synthesis Kit is designed for quick set-up and production of large amounts of RNA in vitro. The
reaction can be set up conveniently by combining the NTP buffer mix, T7 RNA Polymerase mix and a suitable DNA template. The kit also
allows for capped RNA or dye-labeled RNA synthesis by incorporation of cap analog (ARCA, NEB #S1411) or dye-modified nucleotides.
RNA synthesized with the kit can be used for RNA structure and function studies, ribozyme biochemistry, as probes for RNase protection
assays and hybridization based blots, anti-sense RNA and RNAi experiments, microarray analysis and microinjection, as well as in vitrotranslation and RNA vaccines.
To synthesize high specific activity radioactive RNA probes or RNA with 100% substitution of one or more modified nucleotides werecommend using the T7 High Yield RNA Synthesis Kit (NEB #E2040), in which the four nucleotides are supplied separately.

DNA Template Preparation
Linearized plasmid DNA, PCR products or synthetic DNA oligonucleotides can be used as templates for in vitro transcription with the
HiScribe T7 Quick High Yield RNA Synthesis Kit, provided that they contain a double-stranded T7 promoter region upstream of thesequence to be transcribed. Figure 1 illustrates the minimal T7 promoter sequence, as well as a run-off transcript after T7 transcription.
Figure 1. Transcription by T7 RNA Polymerase

Plasmid Templates
It is of the utmost importance to begin the HiScribe T7 Quick High Yield RNA Synthesis Kit with highly purified, completely linearized
plasmid template. Quality of the template DNA affects transcription efficiency, as well as the integrity of the RNA synthesized. Yield iscommensurate with template purity. Any purification method may be used, as long as the product is predominately supercoiled and free ofcontaminating RNase, protein, RNA and salts.
To produce an RNA transcript of defined length, plasmid DNA must be completely linearized with a restriction enzyme, downstream of theinsert to be transcribed. In contrast, circular plasmid templates will generate long heterogeneous RNA transcripts in higher quantities
because of the high processivity of T7 RNA Polymerase. NEB has a large selection of restriction enzymes for this purpose; we recommend
selecting restriction enzymes that generate blunt ends or 5´-overhangs.
After linearization, we recommend purifying the template DNA by phenol:chloroform extraction:
1.

Extract DNA with an equal volume of 1:1 phenol:chloroform mixture, and repeat, if necessary.

2.

Extract twice with an equal volume of chloroform to remove residual phenol.

3.

Precipitate the DNA by adding 1/10th volume of 3 M sodium acetate, pH 5.2, and two volumes of ethanol. Incubate at –20°C for atleast 30 minutes.

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4.

Pellet the DNA in a microcentrifuge for 15 minutes at top speed. Carefully remove the supernatant.

5.

Rinse the pellet by adding 500 μl of 70% ethanol and centrifuging for 15 minutes at top speed. Carefully remove the supernatant.

6.

Air dry the pellet and resuspend it in nuclease-free water at a concentration of 0.5–1 μg/μl.

PCR Templates
PCR products containing a T7 promoter in the correct orientation can be transcribed. While PCR mixture may be used directly, better
yields will be obtained with purified PCR products. PCR products can be purified according to the protocol for plasmid restriction digests
above, or by using commercially available spin columns (we recommend the Monarch PCR & DNA Cleanup Kit, NEB #T1030). PCR
products should be run on an agarose gel to estimate concentration and to confirm amplicon size prior to its use as a template in the T7
Quick RNA transcription reaction. Generally, 0.1–0.5 μg of PCR fragments can be used in a 20 μl in vitro transcription reaction.
Synthetic DNA Oligonucleotides
Synthetic DNA Oligonucleotides, which are either entirely double-stranded or mostly single-stranded, with a double-stranded T7 promoter
sequence can be used in the T7 Quick transcription reaction. In general, the yields are relatively low and also variable depending upon thesequence, purity and preparation of the synthetic oligonucleotides.

RNA Synthesis Protocols
We strongly recommend wearing gloves and using nuclease-free tubes and reagents to avoid RNase contamination. Reactions are typically
20 μl but can be scaled up as needed. Reactions should be assembled in nuclease-free microfuge tubes or PCR strip tubes.
Standard RNA Synthesis
1.

Thaw the necessary kit components, mix and pulse-spin in microfuge to collect solutions to bottom of tubes. Keep on ice.

3.

Assemble the reaction at room temperature in the following order:

REAGENTAMOUNTFINAL
CONCENTRATION

Nuclease-free water

X µl

NTP Buffer Mix

10 µl

10 mM each NTP final

Template DNA

X µl

1 µg

DTT (0.1M)*

1 µl

5 mM

T7 RNA Polymerase Mix

2 µl

Total Reaction Volume

20 µl* Addition of DTT to the reaction is optional but recommended. See Troubleshooting section for details.

3.

Mix thoroughly and pulse-spin in a microfuge. Incubate at 37°C for 2 hours.

Reaction time depends on template amount, quality and RNA transcript length. For reactions with transcripts longer than 0.3 kb, 2 hour
incubation should give you the maximum yield. For reaction times of 60 minutes or less, a water bath or heating block may be used; forreaction times longer than 60 minutes, we recommend using a dry air incubator or a thermocycler to prevent evaporation of the sample.
For reactions with short RNA transcripts (< 0.3 kb), we recommend an incubation time of 4 hours or longer. It is safe to incubate thereaction for 16 hours (overnight). For example, we have achieved good yield with only 0.2 μg plasmid template encoding a 50-mer RNA
by incubating the reaction overnight at 37°C.
Reaction set up for short transcripts (< 0.3 kb):

REAGENTAMOUNTFINAL
CONCENTRATION

Nuclease-free water

X µl

NTP Buffer Mix

10 µl

6.7 mM each NTP final

Template DNA

X µl

1 µg

DTT (0.1M)*

1.5 µl5 mM

T7 RNA Polymerase Mix

2 µl

Total Reaction Volume

30 µl* Addition of DTT to the reaction is optional but recommended. See Troubleshooting section for details.

3Compared to the standard reaction, this reaction uses 10 μl more water. The volume of NTP Buffer Mix and T7 RNA Polymerase Mix,
however, remains the same. The kit contains sufficient materials for 50 reactions.
Note that the amount of NTP Buffer Mix in a standard 20 μl reaction can vary from 2 to 10 μl. The final yield is proportional to the amountof input nucleotides, meaning that the nucleotide incorporation efficiency remains the same when different amounts of NTP are used.
Figure 2 shows the time course of standard RNA synthesis from 1 μg control DNA template coding for a 1.8 kb RNA transcript with the T7
Quick Kit using 10 μl and 5 μl NTP Buffer Mix in a 20 μl reaction.
Figure 2. RNA synthesis with different amounts of NTP

Reactions were incubated at 37°C in a thermocycler. Transcripts
were purified by spin columns and quantified on a NanoDrop™
Spectrophotometer.
4.

Optional: DNase treatment to remove DNA template. Standard reactions are capable of generating large amounts of RNA, atconcentrations up to 10 mg/ml. As a result, the reaction mixture is quite viscous. It is easier to perform DNase treatment after thereaction mixture is diluted. To remove template DNA, add 30 μl nuclease-free water to each 20 μl reaction, followed by 2 μl of DNase
I (RNase-free), mix and incubate for 15 minutes at 37°C.

5.

Proceed with purification of synthesized RNA (we recommend the Monarch RNA Cleanup Kits, NEB #T2040 or #T2050) or analysisof transcription products by gel electrophoresis.

Capped RNA Synthesis
The kit formulation allows for efficient capped RNA synthesis using cap analog (ARCA). The recommended ratio of cap analog to GTP is4:1. Increasing the ratio of cap analog to GTP will increase the proportion of capped RNA transcripts; however, it also significantly
decreases the yield of the reaction. Cap analogs are sold separately. Please refer to the ordering information section or www.neb.com for
more information.
1.

Prepare 40 mM cap analog. Cap analog (ARCA, NEB #S1411) is supplied in a lyophilized form of 1 µmol per tube. Dissolving it in
25 μl nuclease-free water will yield a concentration of 40 mM.

2.

Thaw the necessary kit components, mix and pulse-spin in a microfuge to collect solutions to the bottoms of tubes.

3.

Assemble the reaction at room temperature in the following order:

REAGENTAMOUNTFINAL
CONCENTRATION

Nuclease-free water

X µl

NTP Buffer Mix

2 µl

2 mM each NTP final

Cap Analog (40 mM)

4 µl

8 mM final

Template DNA

X µl

1 µg

DTT (0.1M)*

1 µl

5 mM

T7 RNA Polymerase Mix

2 µl

Total Reaction Volume

20 µl* Addition of DTT to the reaction is optional but recommended. See Troubleshooting section for details.

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4.

Mix thoroughly, pulse-spin and incubate at 37°C for 2 hours.
The yield per reaction is 30–40 μg RNA with approximately 80% capped RNA transcripts. Figure 3 shows the time course of capped
RNA synthesis from 1 µg control template. Most reactions will be complete in 1 hour.
Figure 3. Capped RNA Synthesis with ARCA

Reactions were incubated at 37°C in a thermocycler. Transcripts were purified
by spin columns and quantified on a NanoDrop Spectrophotometer.
5.

Optional: To remove template DNA, add 2 μl of DNase I (RNase-free), mix and incubate at 37°C for 15 minutes.

6.

Proceed with purification of synthesized RNA (we recommend the Monarch RNA Cleanup Kits, NEB #T2040 or #T2050) or analysisof transcription products by gel electrophoresis.

RNA Synthesis with Modified Nucleotides
The kit is capable of synthesizing biotin- or dye-modified RNA with the following protocol. The recommended molar ratio of modified
NTP (Biotin-, Fluorescein-, Digoxigenin-, or Aminoallyl-NTP) to standard NTP is 1:2. The following reaction set-up assumes modified
UTP is used. Please note that Dye- or Biotin-NTPs are not supplied with the kit.
1.

Thaw the necessary kit components, mix and pulse-spin in microfuge to collect solutions to the bottoms of tubes.

2.

Assemble the reaction at room temperature in the following order:
REAGENTAMOUNTFINAL
CONCENTRATION

Nuclease-free water

X µl

NTP Buffer Mix

5 µl

5 mM each NTP final

Modified UTP (10 mM)

5 µl

2.5 mM final

Template DNA

X µl

1 µg

DTT (0.1M)*

1 µl

5 mM

T7 RNA Polymerase Mix

2 µl

Total Reaction Volume

20 µl* Addition of DTT to the reaction is optional but recommended. See Troubleshooting section for details.

3.

Mix thoroughly, pulse-spin and incubate at 37°C for 2 hours. For short (< 300 nt) transcripts incubate at 37°C for 4–16 hours.
Note that the ratio of modified nucleotide to standard nucleotide can be adjusted by varying the amount of the NTP Buffer Mix andmodified nucleotide. For complete modified nucleotide substitution we recommended using the T7 High Yield RNA Synthesis Kit(NEB #E2040), in which the four nucleotides are supplied separately. Figure 4 shows the time course of labeled RNA synthesis using
1 µg control template with Biotin-16-UTP and Fluorescein-12-UTP following the above reaction setup.
Modified ribonucleotides reduce transcription efficiency; therefore, lower transcription yields should be expected as compared totranscription using unmodified NTPs. In general, Biotin-NTP and Aminoallyl-NTP have an insignificant effect on yields, while lower
yields can be expected for transcription reactions containing Fluorescein-NTP or Cy-NTP. In addition, transcripts containing modified
ribonucleotides have reduced electrophoretic mobility due to higher molecular weight.

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Figure 4. Capped RNA Synthesis with ARCA

Reactions were incubated at 37°C in a thermocycler. Transcripts were purified
by spin columns and quantified on a NanoDrop Spectrophotometer.
4.

Optional: To remove template DNA, add 30 μl nuclease-free water and 2 μl of DNase I (RNase-free), mix and incubate at 37°C for
15 minutes

5.

Proceed with purification of synthesized RNA (we recommend the Monarch RNA Cleanup Kits, NEB #T2040 or #T2050) or analysisof transcription products by gel electrophoresis.

Purification of Synthesized RNA
Synthesized RNA can be purified by LiCl precipitation, phenol:chloroform extraction followed by ethanol precipitation, or by using a spincolumn based method. If absolute full length RNA is required, we recommend gel purification.
LiCl Precipitation
The kit includes LiCl solution for quick recovery of the synthesized RNA. LiCl precipitation of RNA is effective in removing the majority
of unincorporated NTPs and enzymes. However, RNAs shorter than 300 bases or at concentrations lower than 0.1 mg/ml do not precipitate
well. In such cases, other purification methods may be used. LiCl purified RNA is suitable for cap addition with NEB’s Vaccinia Capping
System (NEB #M2080) and Poly(A) tailing with NEB’s Poly(A) Polymerase (NEB #M0276).
1.

Adjust the reaction volume to 50 μl by adding nuclease-free water.

2.

Add 25 μl LiCl solution and mix well.

3.

Incubate at –20°C for 30 minutes.

4.

Centrifuge at 4°C for 15 minutes at top speed to pellet the RNA.

5.

Remove the supernatant and rinse the pellet with 500 μl of ice cold 70% ethanol.

6.

Resuspend the RNA in 50 μl of 0.1 mM EDTA. Store the RNA at –20°C or below.

Phenol:chloroform Extraction and Ethanol Precipitation
For removal of proteins and most of the free nucleotides, phenol:chloroform extraction and ethanol precipitation of RNA transcripts is thepreferred method.
1.

Adjust the reaction volume to 180 μl by adding nuclease-free water. Add 20 μl of 3 M sodium acetate, pH 5.2 or 20 μl of 5 M
ammonium acetate and mix thoroughly.

2.

Extract with an equal volume of 1:1 phenol:chloroform mixture, followed by two extractions with chloroform. Collect the aqueous
phase and transfer to a new tube.

3.

Precipitate the RNA by adding 2 volumes of ethanol. Incubate at –20°C for at least 30 minutes and collect the pellet by centrifugation.

4.

Remove the supernatant and rinse the pellet with 500 μl of ice cold 70% ethanol.

5.

Resuspend the RNA in 50 μl 0.1 mM EDTA. Store the RNA at –20°C or below.

Spin Column Purification
Spin columns will remove unincorporated nucleotides, proteins and salts. We recommend the Monarch RNA Cleanup Kits (NEB #T2030,
T2040 or T2050). Adjust the volume of the reaction mixture to 100 μl by adding nuclease-free water and mix well. Purify the RNA byfollowing the spin column manufacturer’s instructions. Each reaction produces up to 200 μg of RNA, which may exceed column capacity,
thus requiring additional columns.

6Gel Purification
When high purity RNA transcript is desired, we recommend gel purification of the transcription product. The Monarch RNA Cleanup Kits
(NEB #T2030, #T2040, #T2050) can be used for RNA gel extraction (see protocol included in NEB #T2030, #T2040, #T2050 productmanual).

Evaluation of Reaction Products
Quantification by UV Light AbsorbanceRNA concentration can be determined by measuring the ultraviolet light absorbance at 260 nm, however, any unincorporated nucleotides
and template DNA in the mixture will affect the reading. Free nucleotides from the transcription reaction must be removed before the RNA
concentration can be quantified. A 1:200 dilution of a sample of the purified RNA should give an absorbance reading in the linear range ofa spectrophotometer. RNA dilution may not be necessary if using a NanoDrop Spectrophotometer. A NanoDrop Spectrophotometer candirectly read RNA concentrations from 10 ng/μl to 3000 ng/μl. For single-stranded RNA, 1 A260 is equivalent to an RNA concentration of
40 μg/ml. The RNA concentration can be calculated as follows:
A260 x dilution factor x 40 = __ μg/ml RNA
Analysis of Transcription Products by Gel Electrophoresis
To evaluate transcript length, integrity and quantity, an aliquot of the transcription reaction should be run on an appropriate denaturing
agarose or polyacrylamide gel. Transcripts larger than 0.3 kb can be run on agarose gels, whereas denaturing polyacrylamide gels (5–15%)
are necessary for smaller transcripts. The gels should be run under denaturing conditions to minimize formation of secondary structures by
the transcript.
1.

Preparation of denaturing gelsa. Denaturing agarose gel:To make a 100 ml 1% denaturing agarose gel, add 1 gram agarose powder to 72 ml nuclease-free water. Melt the agarose and add
10 ml 10X MOPS buffer. Then, in a fume hood, add 18 ml fresh formaldehyde (37%), mix well. Pour the gel.
10X MOPS gel running buffer: 0.4 M MOPS (pH 7.0), 0.1 M Sodium
Acetate, 10 mM EDTA
b. Denaturing PAGE/Urea Gel:
5–15% PAGE/Urea gel. We recommend using commercially available premade gels. Use standard TBE gel running buffer.
10X TBE buffer: 0.9 M Tris Base, 0.9 M Boric Acid, 20 mM EDTA.

2.

Gel electrophoresis of non-radiolabeled RNA
a. Mix 0.2–1 μg RNA sample with an equal volume of RNA Loading Dye (2X, NEB #B0363).
b. Denature the RNA sample and an aliquot of RNA marker by heating at 65–70°C for 5–10 minutes.
c. Pulse-spin prior to loading onto gel.
d. Visualizing RNA by staining the gel with SYBR® Gold or ethidium bromide.

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Troubleshooting
Control Reaction
The FLuc control template DNA is a linearized plasmid containing the firefly luciferase gene under the transcriptional control of the T7
promoter. The size of the runoff transcript is 1.8 kb. The control reaction should yield  150 μg RNA transcript in 2 hours.
If the control reaction is not working, there may be technical problems during reaction set up. Repeat the reaction by following the protocol
carefully; take all precautions to avoid RNase contamination. Contact NEB for technical assistance.
The control plasmid sequence can be found within the DNA Sequences and Maps Tool under the name “FLuc Control Plasmid”. The FLuccontrol template is generated by linearizing the plasmid with StuI.
Low Yield of Full-length RNA
If the transcription reaction with your template generates full-length RNA, but the yield is significantly lower than expected, it is possiblethat contaminants in the DNA template are inhibiting the RNA polymerase, or the DNA concentration may be incorrect. Alternatively,
additional purification of DNA template may be required. Phenol:chloroform extraction is recommended (see template DNA preparation
section).
Addition of DTT
Addition of DTT (5 mM final) to the reaction is optional but recommended. The RNA polymerase in the kit is sensitive to oxidation andcould result in lower RNA yield over time due to repeated handling etc. Adding DTT to the reaction may help restore the kit performancein such cases. Adding DTT will not compromise the reaction in any situation.
Low Yield of Short Transcript
High yields of short transcripts (< 0.3 kb) are achieved by extending incubation time and increasing the amount of template. Incubation ofreactions up to 16 hours (overnight) or using up to 2 μg of template will help to achieve maximum yield.
RNA Transcript Smearing on Denaturing Gel
If the RNA appears degraded (e.g., smeared) on denaturing agarose or polyacrylamide gel, the DNA template is likely contaminated with
RNase. DNA templates contaminated with RNase can affect the length and yield of RNA synthesized (a smear below the expectedtranscript length). If the plasmid DNA template is contaminated with RNase, perform phenol:chloroform extraction, then ethanol
precipitate and dissolve the DNA in nuclease-free water (see template DNA preparation section).
RNA Transcript of Larger Size than Expected
If the RNA transcript appears larger than expected on a denaturing gel, template plasmid DNA may be incompletely digested. Even small
amounts of undigested circular DNA can produce large amounts of long transcripts. Check template for complete digestion. If undigested
plasmid is confirmed, repeat restriction enzyme digestion.
Larger size bands may also be observed when the RNA transcript is not completely denatured due to the presence of strong secondary
structures.
RNA Transcript of Smaller Size than Expected
If denaturing gel analysis shows the presence of smaller bands than the expected size, it is most likely due to premature termination by
the polymerase. Some sequences with resemblance to T7 RNA Polymerase termination signals will cause premature termination.
Incubating the transcription reaction at lower temperatures, for example at 30°C, may increase the proportion of full-length transcript,
however the yield will be decreased. For GC rich templates, or templates with secondary structures, incubation at 42°C may improveyield of full-length transcript.

8Ordering Information
NEB #PRODUCT

SIZE

E2050S/L

HiScribe T7 Quick High Yield RNA Synthesis Kit

50/250 reactions

COMPANION PRODUCTS
NEB #PRODUCT

SIZE

T2040S/L

Monarch RNA Cleanup Kit (50 µg)

10/100 preps

T2030S/L

Monarch RNA Cleanup Kit (10 µg)

10/100 preps

T2050S/L

Monarch RNA Cleanup Kit (500 µg)

10/100 preps

B0363S

RNA Loading Dye (2X)

4 x 1 ml

M0307S/L

RNase Inhibitor, Human Placenta

2,000/10,000 units

M0314S/L

RNase Inhibitor, Murine

3,000/15,000 units

M0303S/L

DNase I (RNase-Free)

1,000/5,000 units

M0493S/LQ5 Hot Start High-Fidelity DNA Polymerase

100/500 units

N0362S

ssRNA Ladder

25 gel lanes

N0364S

Low Range ssRNA Ladder

25 gel lanes

S1411S/L

3´-O-Me-m7G(5´)ppp(5´)G RNA Cap Structure Analog

1/5 µmol

S1405S/L

m7G(5´)ppp(5´)A RNA Cap Structure Analog

1/5 µmol

S1406S/LG(5´)ppp(5´)A RNA Cap Structure Analog

1/5 µmol

S1407S/LG(5´)ppp(5´)G RNA Cap Structure Analog

1/5 µmol

S1404S/L

m7G(5´)ppp(5´)G RNA Cap Structure Analog

1/5 µmol

E3320S

RNase Contamination Assay Kit

50 reactions

M2080S

Vaccinia Capping System

400 units

M0366SmRNA Cap 2´-O-Methyltransferase

2,000 units

M0276S/L

E. coli Poly(A) Polymerase

100/500 units

N0466S/L

Ribonucleotide Solution Mix

10/50 µmol of eachN0450S/L

Ribonucleotide Solution Set

10/50 µmol of each9

Revision History
REVISION #1.0

DESCRIPTION
N/A

DATE
2/13

1.1

5/13

2.0

4/14

2.1

1/17

3.0

1/19

4.0

Applied new manual format.

4/20

5.0

Updated to included addition of DTT. Also updated locationof control plasmid and LiCl Solution concentration. Also
updated table formatting and legal footer.

7/23

6.0

Updated to include new L size.

1/24

6.1

Updated final concentration of NTP Buffer Mix in table on
page 5 to “5 mM each NTP final”.

9/24

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
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does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. See
www.neb.com/trademarks. The use of these products may require you to obtain additional third-party intellectual property rights for certain applications. For more information,please email busdev@neb.com.
B CORPORATION® is a registered trademark of B Lab IP, LLC, Inc.
NANODROP® is a registered trademark of NanoDrop Technologies, Inc.
SYBR® is a registered trademark of Life Technologies, Inc.
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