pEASY ®- T3 Cloning Vector provides dual EcoR I and dual Not I enzyme sites. It is designed for cloning and sequencing Taq-amplified PCR products. The cloned insert can be released from a single enzyme digestion.

•5 minutes fast ligation of Taq-amplified PCR products.

•Ampicillin resistance genes for selection.

•Easy blue/white selection.

•T7 promoter, SP6 promoter, M13 forward and M13 reverse primers for sequencing.

•T7 promoter and SP6 promoter for in vitro transcription.

•Trans1-T1 Phage Pesistant Chemically Competent Cells, high transformation efficiency (>109 cfu/μg pUC19 DNA) and fast growing.



1.Primer requirement: primer cannot be phosphorylated

2.PCR Enzyme: Taq DNA polymerases

3.Reaction conditions: in order to ensure the integrity of amplification products, 5-10 minutes of post-extension step is required. After amplification reaction, use agarose gel electrophoresis to verify the quality and quantity of PCR product


Reaction System

Add following components into a microcentrifuge tube.

PCR products 0.5-4 μl (can be increased or reduced based on PCR product yield, no more than 4 μl)

pEASY ®- T3 Cloning Vector 1 μl

Gently mix well, incubate at room temperature (20oC-37oC) for 5 minutes. After reaction, place the tube on ice.

1. Optimal amount of insert

Molar ratio of vector to insert = 1:7 (1 kb, ~20 ng; 2 kb, ~40 ng)

2.Optimal volume of vector: 1 μl(10 ng)

3.Optimal reaction volume: 3~5 μl

4.Optimal incubation time

(1)0.1~1 kb (including 1 kb): 5~10 minutes

(2)1~2 kb (including 2 kb): 10~15 minutes

(3)2~3 kb (including 3 kb): 15~20 minutes

(4)≥3 kb: 20~30 minutes

Use the maximum incubation time if the insert is gel purified.

5.Optimal incubation temperature: for most PCR inserts, the optimal temperature is about 25oC; for some PCR inserts, optimal results can be achieved with higher temperature (up to 37oC).



1.Add the ligated products to 50 μl of Trans1-T1 Phage Resistant Chemically Competent Cell and mix gently (do not mix by pipetting up and down).

2.Incubate on ice for 20~30 minutes.

3.Heat-shock the cells at 42oC for 30 seconds.

4.Immediately place the tube on ice for 2 minutes.

5.Add 250 μl of room temperature SOC or LB medium. Shake the tube at 37oC (200 rpm) for 1 hour.

6.In the meantime, mix 8 μl of 500 mM IPTG with 40 μl of 20 mg/ml X-gal. Spread them evenly onto a selective LB plate. Place the plate at 37oC for 30 minutes.

7.Spread 200 μl or all transformants on the pre-warmed plate. Incubate at 37oC overnight.


Contents& storage





(20 rxns)

(60 rxns)


pEASY ®-T3 Cloning Vector (10 ng/μl)

20 µl

3×20 µl

Control Template (5 ng/μl)

5 µl

5 µl

Control Primers (10 μM)

5 µl

5 µl

M13 Forward Primer (10 μM)

50 µl

150 µl

M13 Reverse Primer (10 μM)

50 µl

150 µl

Trans1-T1 Phage Resistant

Chemically Competent Cell

10×100 μl

30×100 μl


Citations & references
Evolution and Expression Plasticity of Opsin Genes in a Fig Pollinator, Ceratosolen solmsiPLoS One3.73Wang B. et al.2013http://www.ncbi.nlm.nih.gov/pubmed/23342036
The impact of dissolved organic carbon on the spatial variability of methanogenic archaea communities in natural wetland ecosystems across ChinaApplied Microbiology and Biotechnology3.689Liu D, et al.2012 Octhttp://www.ncbi.nlm.nih.gov/pubmed/22218772
Arsenic induces functional re-expression of estrogen receptor α by demethylation of DNA in estrogen receptor-negative human breast cancerPLoS One3.73Du J, et al.2012http://www.ncbi.nlm.nih.gov/pubmed/22558281
Complex evolution in Arundinarieae (Poaceae: Bambusoideae): incongruence between plastid and nuclear GBSSI gene phylogeniesMolecular Phylogenetics and Evolution4.066Zhang YX, et al.2012 Junhttp://www.ncbi.nlm.nih.gov/pubmed/22415014
DsbM, a novel disulfide oxidoreductase affects aminoglycoside resistance in Pseudomonas aeruginosa by OxyR-regulated responseJournal of Microbiology1.276Wang X, et al.2012 Dechttp://link.springer.com/article/10.1007/s12275-012-2177-3
Sinomyces: a new genus of anamorphic PleosporaceaeFungal Biology2.082Wang Y, et al.2011 Febhttp://www.ncbi.nlm.nih.gov/pubmed/21315316
Identification of ethylene-responsive genes in ethrel-treated shoot apices of cucumber by suppression subtractive hybridizationPlant Molecular Biology Reporter5.319Jin X, et al.2011 Dec


Three novel species of Stemphylium from Sinkiang, China: their morphological and molecular characterizationMycological Progress1.606Pei YF, et al.2011 Mayhttp://link.springer.com/article/10.1007/s11557-010-0686-1
Molecular and morphological description of two new species of Stemphylium from China and FranceMycologia2.11Wang Y, et al.2010 May-Junhttp://www.ncbi.nlm.nih.gov/pubmed/20524602
Ulocladium cantlous sp. nov. isolated from northwestern China: its morphology and molecular phylogenetic positionMycologia2.11Wang Y, et al.2010 Mar-Aprhttp://www.ncbi.nlm.nih.gov/pubmed/20361504
Molecular characterization, immunohistochemical localization and expression of a ribosomal protein L17 gene from Apis cerana ceranaInsect Biochemistry and Physiology1.515Fei Meng, et al.2010 Octhttp://onlinelibrary.wiley.com/doi/10.1002/arch.20386/abstract
The complete mitochondrial genome of Spilonota lechriaspis Meyrick (Lepidoptera: Tortricidae)Molecular Biology Reports2.506Jin-Liang Zhao, et al.2011 Aughttp://link.springer.com/article/10.1007/s11033-010-0491-6
Halomonas songnenensis sp. nov., a moderately halophilic bacterium isolated from saline and alkaline soilsIJSEM Juquan Jiang,et al.2014 Mayhttp://ijs.sgmjournals.org/content/64/Pt_5/1662.short
Characterization of a mitochondrial manganese superoxide dismutase gene from Apis cerana cerana and its role in oxidative stressJournal of Insect Physiology2.379Haihong Jia, et al.2014 Janhttp://www.sciencedirect.com/science/article/pii/S002219101300231X
Cryptic diversity in host-associated populations of Tetra pinnatifidae (Acari: Eriophyoidea): What do morphometric, mitochondrial and nuclear data reveal and conceal?Bulletin of Entomological Research1.987Hao-Sen Li, et al.2014 Apr



Identification of Rhizoctonia solani AG1-IB as a causal agent of leaf blight in Cynanchum paniculatum in ChinaJournal of General Plant Pathology0.893Ai Xin Wang, et al.2014 Janhttp://link.springer.com/article/10.1007/s10327-013-0489-8
MOLECULAR CHARACTERIZATION OF SOLUBLE AND MEMBRANE-BOUND TREHALASES IN THE COTTON MIRID BUG, Apolygus lucorumArchives of Insect Biochemistry and Physiology1.515Yongan Tan, et al.2014 Jun



RNAi mediated knockdown of the ryanodine receptor gene decreases chlorantraniliprole susceptibility in Sogatella furciferaPesticide Biochemistry and Physiology2.111Yao Yang, et al.2014 Janhttp://www.sciencedirect.com/science/article/pii/S0048357513001922
Pollen and seed transmission of Cucumber green mottle mosaic virus in cucumberPlant Pathology2.729H.W.Liu, et al.2014 Feb


Biochemical functions of the glutathione transferase supergene family of Larix kaempferiPlant Physiology and Biochemistry2.775Qi Yang, et al.2014 Aprhttp://www.sciencedirect.com/science/article/pii/S0981942814000473
RNA interference-mediated silencing of a Halloween gene spookier affects nymph performance in the small brown planthopper Laodelphax striatellusInsect Science1.786Shuang Jia, et al.2014 Apr




Differential Expression Patterns in Chemosensory and Non-Chemosensory Tissues of Putative Chemosensory
Genes Identified by Transcriptome Analysis of Insect Pest the Purple Stem Borer Sesamia inferens (Walker)
PLOS one3.7Zhang, YN,et al.2013 Aughttp://journals.plos.org/plosone/article?id=10.1371/journal.pone.0069715
A novel bifunctional pectinase from Penicilliumoxalicum SX6with separate pectin methylesterase and polygalacturonasecatalytic domainsAppl Microbiol Biotechnol3.689Tao Tu, et al.2014 Janhttp://link.springer.com/article/10.1007/s00253-014-5533-x
Two Origins of Blastemal Progenitors Define Blastemal
Regeneration of Zebrafish Lower Jaw
PloSOne3.54Xuelong Wang, et al.2014 Dechttp://journals.plos.org/plosone/article?id=10.1371/journal.pone.0045380
Two Metallothionein Genes in Oxya chinensis: Molecular
Characteristics, Expression Patterns and Roles in Heavy
Metal Stress
PloSOne3.54Yaoming Liu, et al.2014 Novhttp://journals.plos.org/plosone/article?id=10.1371/journal.pone.0112759
High Level Expression of a Novel Family 3 Neutral β-Xylosidase from Humicola insolens Y1 with High Tolerance to D-XylosePloSOne3.54Wei Xia, et al.2015 Febhttp://journals.plos.org/plosone/article?id=10.1371/journal.pone.0117578
Electrical signals generated by soil microorganisms in microbial fuel cells respond linearly to soil Cd 2+ pollutionGeoderma2.772Yun-Bin Jiang, et al.2015 AprJiangsu Provincial Key Laboratory of Materials Cycling and Pollutio

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pEASY®-T3 Cloning Kit

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  • Product Code: CT301
  • Availability: In Stock
  • $255.00

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