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BigEasy v2.0 Cloning Kits with pJAZZ Linear Notl Vectors

BigEasy v2.0 Linear Cloning Kits include:  Dephosphorylated pJAZZ vector pre-cut at either a SmaI (blunt) or NotI site, CloneSmart DNA Ligase, CloneDirect™ 10X Ligation Buffer (includes ATP), Sequencing Primers, Positive Control Insert DNA, DNA Terminator® End Repair Enzyme, BigEasy-TSA Electrocompetent Cells in SOLO packaging (1 transformation per tube), Recovery Medium, Transformation Control DNA, and complete protocols. BigEasy-TSA Electrocompetent Cells are also available separately.

BigEasy v2.0 Cloning Kits with pJAZZ Linear Notl Vectors

Clone the un-clonable, including large or highly repetitive DNA sequences, with this unique linear cloning system.
  • Efficiently clone any insert up to 30 kb.
  • Create libraries from even A/T or G/C-rich genomes.
  • Clone gene clusters or operons.
  • Maximum insert stability.
  • Inducible copy number.
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Product information

The BigEasy Kit with the pJAZZ vector is ideal for constructing bias-free, large-insert genomic libraries, or for cloning difficult DNA of any size up to 30 kb. Because the novel pJAZZ vector is maintained as a linear molecule, the ends of the vector can rotate freely. As a result, the vector does not supercoil. Without the torsional stress induced by supercoiling, otherwise unclonable sequences (e.g., repetitive, or A/T-rich or G/C-rich sequences) are stabilised. The pJAZZ linear cloning vector is maintained at low copy number (5-10/cell) in BigEasy-TSA™ Electrocompetent Cells (>4 × 1010 cfu/µg), which are required for transformation and propagation. The copy number can be induced 5-10X in this strain. In addition, the pJAZZ vector incorporates CloneSmart® technology for transcription-free cloning, which further increases insert stability. pJAZZ clones can be isolated using conventional plasmid prep methods and sequenced using standard techniques.

pJAZZ-OC and pJAZZ-OK vectors

Figure 1. pJAZZ-OC and pJAZZ-OK linear vectors. RepA, replication factor and low copy origin of replication (~2-4 per cell; inducible 5-10 fold); Camr - chloramphenicol resistance gene; Kanr - kanamycin resistant gene; telN - protelomerase gene; cB - replication regulator. Approximate positions of transcription terminators (T) are indicated.

Figure 2. Sequence trace of a Piromyces clone, showing extremely high (96%) AT content (see figure 6).

Figure 3. 15-20 kb PCR amplification products cloned into the pJAZZ® linear vector.

Figure 4. Mouse genomic DNA sheared to
6-20 kb and cloned into the pJAZZ linear vector.

Figure 5. Oxytricha trifallax genomic DNA (75-85% AT) sheared to 6-20 kb and cloned into the pJAZZ linear vector.

Figure 6. Piromyces genomic DNA (85% AT) cloned in the pJAZZ vector. This DNA was unclonable in all other vectors.

Convenient Success

Efficiently clone the most difficult DNAs

The BigEasy v2.0 Kits eliminate tedious vector and competent cell preparation, as well as time-consuming QC testing. Kits include optimised reagents, ligation-ready vector, highly efficient electrocompetent cells, detailed instructions, and trouble-shooting guides.

Published Paper:

Godiska R, Mead DA, Dhodda V, Hochstein R, Karsi A, Usdin K, Entezam A, Ravin N. (2010) Linear plasmid for cloning large or repetitive sequences in E. coli. Nuc. Acids Res. 38:e88. Read Paper >>

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