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CloneSmart Blunt End Cloning Kits for Difficult To Clone DNA

Successfully clone difficult DNAs like AT-rich regions or toxic genes
• Clone difficult DNAs like AT-rich regions or toxic genes
• Benefit from transcription- and translation-free cloning and increased insert stability
• Eliminate screening of recombinants due to <1% background
• Choose between ampicillin or kanamycin selection
• Achieve high cloning efficiencies with electrocompetent or chemically competent cells

The transcription- and translation-free pSMART vectors contained in the CloneSmart Blunt Cloning Kits minimize many of the problems associated with cloning recalcitrant DNA into conventional plasmid vectors. Vector-driven transcription and translation of the insert DNA is completely eliminated, as these vectors do not contain a promoter or indicator gene that may initiate transcription through the cloning site. Moreover, insert-driven transcription into the vector is blocked by strong terminators flanking the cloning site, enabling the successful cloning of inserts containing coding regions, strong promoters, short repeats, or incompatible secondary structures.
The low copy number versions of pSMART further increase the chances of cloning intact sequences that are difficult to maintain in typical vectors.
For example, shotgun cloning of an AT-rich genome such as Lactobacillus helveticus in pSMART resulted in 30 times more stable clones than in pUC19. A number of additional difficult-to-clone genes and genomes have been obtained with this technology, see product citations link below.

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High copy (HC) and low copy (LC) versions of the pSMART transcription-free cloning vectors.
Amp, ampicillin resistance gene; Kan, kanamycin resistance gene; Ori, origin of replication; ROP, "Repressor of Primer" copy number gene.


Background-free Cloning
The pSMART vectors are supplied pre-digested, with blunt dephosphorylated ends, and are qualified to produce 99.5% recombinant clones in typical experiments. The ultra-low background of empty vector (less than 0.5%) eliminates the need to screen for recombinants and enables library construction from nanogram amounts of DNA. Because no screening is required, CloneSmart technology eliminates the need for XGAL/IPTG and removes the uncertainty of false negatives (light blue pUC colonies) and false positives (white colonies that lack inserts).
The small size of the pSMART vectors (1.7-2.0 kb) facilitates subcloning, mutagenesis and transposition experiments.

Selection Guide
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CloneSmart Blunt Cloning Kit Contents
- Vector Premix
- CloneSmart DNA Ligase
- Positive Control Insert DNA
- Sequencing Primers

Lucigen´s highest efficiency competent cells - see link below - ideally fit to the CloneSmart cloning kits.

Also the CloneSmart kits harmonize perfectly with cloning blunt ended PCR fragments, generated by proofreading polymerases such as PRECISOR High-Fidelity DNA Polymerase - see link below.

Related Links

Product Citations
Chemically Competent & Electrocompetent Cells
PRECISOR High-Fidelity DNA Polymerase
NGS Library Prep Kits for Metagenomics


PDF-Downloads - Will open in new browser window

Example: Cloning of AT-rich Lactobacillus helveticus genome

 

 

Description Cat# Size Price    
CloneSmart HCAmp Blunt Cloning Kit (without cells) 40041-2-LU 20 rxns 338 € DETAILS   Add to Cart 
CloneSmart HCAmp Blunt Cloning Kit (without cells) 40041-4-LU 40 rxns 540 € DETAILS   Add to Cart 
CloneSmart HCKan Blunt Cloning Kit (without cells) 40704-2-LU 20 rxns 338 € DETAILS   Add to Cart 
CloneSmart HCKan Blunt Cloning Kit (without cells) 40704-4-LU 40 rxns 540 € DETAILS   Add to Cart 
CloneSmart LCAmp Blunt Cloning Kit (without cells) 40300-2-LU 20 rxns 387 € DETAILS   Add to Cart 
CloneSmart LCAmp Blunt Cloning Kit (without cells) 40300-4-LU 40 rxns 623 € DETAILS   Add to Cart 
CloneSmart LCKan Blunt Cloning Kit (without cells) 40821-2-LU 20 rxns 387 € DETAILS   Add to Cart 
CloneSmart LCKan Blunt Cloning Kit (without cells) 40821-4-LU 40 rxns 623 € DETAILS   Add to Cart 

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