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Pro Ligation-Free Cloning Kit (20 reactions

Cat. No.
E086
Unit
20 Reactions
Cat. No. E086
Name Pro Ligation-Free Cloning Kit (20 reactions)
Unit 20 Reactions
Category Molecular Biology Enzymes and Kits
Description The Pro Ligation-Free Cloning Kit allows for the robust assembly of multiple DNA fragments in one step. Using a ligation-independent cloning method, as many as six DNA fragments are assembled seamlessly without concern for restriction enzyme site availability. abm’s Pro Ligation-Free cloning is a simple yet highly versatile method to rapidly clone highly complex, multiple component constructs.

Key Kit Features:

  • One-step cloning: No more time-consuming, multi-step traditional cloning that depnds on available restrict sites or compatible ends
  • Scar-free cloning: No unwanted/extra base pairs in the inserts
  • Easy-to-customize: Clone in promoter, gene, tag, multi-fragment or whiole vector constructs in one step
  • Ready for downstream applications: Ligase-free reaction, no gel extraction or column purification steps needed
  • Robust assembly: Better performance for difficult, multi-fragment assemblies such as CRISPR multiplex sgRNA vector assembly
  • Increased thermostability: High colony yields

Testimonial:

"I tried the molecular cloning of inserting two DNA fragments (2Kb & 0.5Kb) into a 4Kb backbone by using the kit. After the transformation, I got about 100 colonies, and when I picked 24 colonies for colony PCR, the bright band from gel running told me that I got 20 potential hits, which is awesome. I sent some plasmids for sequencing, the result I received just now unveiled they are carrying the exact right fragment insertion. Collectively, this is a powerful kit."

Jianfeng Lan, The Buck Institute

Application
  • Rapid multi-fragment cloning and assembly of whole vector constructs
  • Clone in a promoter, gene, and tag, all in one step
  • Rapid assembly of CRISPR multiplex gRNA vectors in one ste
Storage Condition Store at -20°C
Search CoA here

Do I need to purify the PCR product first before use with this kit? Or can I use low melting agarose for the PCR gel and just melt the band portion and use in the kit?

The kit can be used in either scenario.

1. The kit comes with Cloning Optimizer and Assembly Enhancer enzymes which allow the user to not have to do gel extraction/purification following the amplification of DNA fragments, as methylated template DNA and single stranded DNA (e.g. primers) will be removed.

2. If PCR reactions contain multiple non-specific bands, it is highly advisable to either optimize the PCR or perform gel extraction, as the contaminating bands will be present in the cloning reaction if not prior removed.

  • Huang, R., Ding, R., Liu, Y., Li, F., Zhang, Z., & Wang, S. A. 2022. GATA transcription factor WC2 regulates the biosynthesis of astaxanthin in yeast Xanthophyllomyces dendrorhous. Microbial Biotechnology, 15(10), 2578-2593.
  • Hwang, B.-J., Gonzales, R., Corzine, S., Stenson, E., Pidugu, L., & Lu, A.-L. (2022). DNA binding by the Rad9A subunit of the Rad9-Rad1-Hus1 complex. PLOS ONE, 17(8), e0272645. https://doi.org/10.1371/journal.pone.0272645
  • Li, L., Liu, W., Fan, N., Li, F., Huang, B., Liu, Q., ... & Wang, X. (2022). Scallop IKK1 Responds to Bacterial and Virus-Related Pathogen Stimulation and Interacts With MyD88 Adaptor of Toll-Like Receptor Pathway Signaling. Frontiers in Immunology, 13. https://doi.org/10.3389/fimmu.2022.869845
  • Huang, B., Wu, Y., Ma, J., Yang, B., Sang, X., Chen, J., Liu, W., Li, F., Li, L., Wang, X., Dong, J., & Wang, X. (2022). The first identified invertebrate LGP2-like homolog gene in the Pacific oyster Crassostrea gigas. Fish & Shellfish Immunology, 128, 238–245. https://doi.org/10.1016/j.fsi.2022.08.005
  • Sang, X., Liu, W., Li, F., Huang, B., Li, L., Wang, X., Dong, J., Ma, J., Chen, J., & Wang, X. (2022). Scallop RIG-I-like receptor 1 responses to polyinosinic:polycytidylic acid challenge and its interactions with the mitochondrial antiviral signaling protein. Fish & Shellfish Immunology, 124, 490–496. https://doi.org/10.1016/j.fsi.2022.04.042
  • Li M., et al. 2022. A Chitin-Binding Protein Ultra-highly expressed in the Outer Fold of Mantle is Related to Shell Colour in Pacific Oyster Crassostrea gigas. https://doi.org/10.55277/researchhub.jg604pn4

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