Cloning
Hieff Clone™ Universal One Step Cloning Kit - 10922ES
A simple, fast, and highly efficient cloning technology and enables directional insertion of any amplified DNA product into any linearized vector at any site. Firstly, the vector is linearized at the cloning site. A small sequence overlapped with each end of the cloning site is added onto the insert through PCR. The insert and the linearized vector, with overlapped sequences of 15 bp - 20 bp on both 5' - and 3' -end, respectively, are mixed in an appropriate ratio and incubated with recombinase at 50℃ for 5-15 min.
Hieff Clone™ Universal one step cloning kit is a novel cloning Kit, independent of DNA ligase, significantly reducing the self-ligated colonies and bringing a true positive rate >95%. For Quick Clone application, it allows multiple overlapping DNA fragments to be seamlessly linked in a one-step, 15–60 minute isothermal reaction. Figure 1 Multi-segment linkage: performance comparison with competitive strains (plate colony). Top half of the picture: 5 fragments+vector; Total segment length: 4000 bp; Vector length: 5000 bp. Bottom half of the picture: 6 fragments+vector; Total segment length: 5000 bp; Vector length: 5000 bp Figure 2 Multi-segment linkage: performance comparison with competitive strains (colony PCR). Top half of the picture: 5 fragments+vector; Total segment length: 4000 bp; Vector length: 5000 bp. Bottom half of the picture: 6 fragments+vector; Total segment length: 5000 bp; Vector length: 5000 bp Selected citations: [1] Multicoloured fluorescent indicators for live-cell and in vivo imaging of inorganic mercury dynamics. Free Radic Biol Med. 2018;121:26-37. doi:10.1016/j.freeradbiomed.2018.04.562(IF:6.020) [2] Characterization of an L-Arabinose Isomerase from Bacillus velezensis and Its Application for L-Ribulose and L-Ribose Biosynthesis. Appl Biochem Biotechnol. 2020;192(3):935-951. doi:10.1007/s12010-020-03380-0(IF:2.277)
Hieff Clone™ Universal one step cloning kit is a novel cloning Kit, independent of DNA ligase, significantly reducing the self-ligated colonies and bringing a true positive rate >95%. For Quick Clone application, it allows multiple overlapping DNA fragments to be seamlessly linked in a one-step, 15–60 minute isothermal reaction. Figure 1 Multi-segment linkage: performance comparison with competitive strains (plate colony). Top half of the picture: 5 fragments+vector; Total segment length: 4000 bp; Vector length: 5000 bp. Bottom half of the picture: 6 fragments+vector; Total segment length: 5000 bp; Vector length: 5000 bp Figure 2 Multi-segment linkage: performance comparison with competitive strains (colony PCR). Top half of the picture: 5 fragments+vector; Total segment length: 4000 bp; Vector length: 5000 bp. Bottom half of the picture: 6 fragments+vector; Total segment length: 5000 bp; Vector length: 5000 bp Selected citations: [1] Multicoloured fluorescent indicators for live-cell and in vivo imaging of inorganic mercury dynamics. Free Radic Biol Med. 2018;121:26-37. doi:10.1016/j.freeradbiomed.2018.04.562(IF:6.020) [2] Characterization of an L-Arabinose Isomerase from Bacillus velezensis and Its Application for L-Ribulose and L-Ribose Biosynthesis. Appl Biochem Biotechnol. 2020;192(3):935-951. doi:10.1007/s12010-020-03380-0(IF:2.277)
Features:
- Simple: Seamlessly assemble and clone up to six DNA fragments in a single reaction
- Flexible: Design guidelines allow assembly into any vector of your choice
- Efficient: Efficient for ligation of one to six fragments
Hieff Clone ™ Universal II One Step Cloning Kit - 10923ES
Hieff CloneTM Universal II One Step Cloning Kit is a new generation homologous recombinant cloning kit. The carefully optimized 2nd generation 2× Hieff CloneTM Universal II Enzyme Premix combines the recombinant enzyme and the buffer required for the recombinant reaction with the addition of a unique recombinant enhancer to significantly improve the efficiency of recombinant cloning.
The kit can be directed to clone PCR products to any site of any vector, compatible with unpurified PCR products, directly recovered PCR products, low concentration of rubber recovered products, this product can reassemble the homologous arm GC content of 30%-70% of the joint fragment. The vector was completely linearized, and homologous sequences of the end of the linearized vector of 15-25 bp were introduced into the 5 'end of the positive and reverse PCR primers of the inserted fragment, so that the 5' and 3 'ends of the PCR products of the inserted fragment had exactly the same sequence corresponding to the two ends of the linearized vector, respectively. Under the action of recombinant enzyme, the recombinant reaction of PCR product and linearized vector can be completed in as little as 5 min at 50℃. The positive rate of cloning can reach more than 95%. Suitable applications: - Rapid Cloning: It allows multiple overlapping DNA fragments to be seamlessly linked in a one-step, 5–50 minute isothermal reaction. - Directed Cloning; Site-Directed Mutagenesis. Figure (Top A-C). A total of 10 ng low-input single fragment was cloned into a 10 kb plasmid. The results showed that 10923ES had better performance than competing products, with high connection efficiency, more colonies and a positive rate of 100%. A-B: Colony plate. The molar ratio of carrier (10 kb) to insert fragment (1 kb) was 1:2. C: Electrophoresis analysis of the insertion fragment, M: YEASEN 10505ES. Plasmid input 40 ng/6 μL, 50°C, 15 mins. Figure (Bottom A-B). A 4.5kb fragment was cloned into a 21 kb plasmid. A: Colony plate. The molar ratio of plasmid to insert fragment was 1:2. B: Electrophoresis analysis of the insertion fragment, M: YEASEN 10505ES. Plasmid input：200 ng/20 μL, 50°C, 40 mins. Selected citations: [1] Reversible phase separation of HSF1 is required for an acute transcriptional response during heat shock. Nat Cell Biol. 2022;24(3):340-352. (IF:28.824) [2] A Selective Small-Molecule c-Myc Degrader Potently Regresses Lethal c-Myc Overexpressing Tumors. Adv Sci (Weinh). 2022;9(8):e2104344. (IF:16.806) [3] Phosphatidic acid suppresses autophagy through competitive inhibition by binding GAPC (glyceraldehyde-3-phosphate dehydrogenase) and PGK (phosphoglycerate kinase) proteins [published online ahead of print, 2022 Mar 15]. Autophagy. 2022;1-15. (IF:16.016)
The kit can be directed to clone PCR products to any site of any vector, compatible with unpurified PCR products, directly recovered PCR products, low concentration of rubber recovered products, this product can reassemble the homologous arm GC content of 30%-70% of the joint fragment. The vector was completely linearized, and homologous sequences of the end of the linearized vector of 15-25 bp were introduced into the 5 'end of the positive and reverse PCR primers of the inserted fragment, so that the 5' and 3 'ends of the PCR products of the inserted fragment had exactly the same sequence corresponding to the two ends of the linearized vector, respectively. Under the action of recombinant enzyme, the recombinant reaction of PCR product and linearized vector can be completed in as little as 5 min at 50℃. The positive rate of cloning can reach more than 95%. Suitable applications: - Rapid Cloning: It allows multiple overlapping DNA fragments to be seamlessly linked in a one-step, 5–50 minute isothermal reaction. - Directed Cloning; Site-Directed Mutagenesis. Figure (Top A-C). A total of 10 ng low-input single fragment was cloned into a 10 kb plasmid. The results showed that 10923ES had better performance than competing products, with high connection efficiency, more colonies and a positive rate of 100%. A-B: Colony plate. The molar ratio of carrier (10 kb) to insert fragment (1 kb) was 1:2. C: Electrophoresis analysis of the insertion fragment, M: YEASEN 10505ES. Plasmid input 40 ng/6 μL, 50°C, 15 mins. Figure (Bottom A-B). A 4.5kb fragment was cloned into a 21 kb plasmid. A: Colony plate. The molar ratio of plasmid to insert fragment was 1:2. B: Electrophoresis analysis of the insertion fragment, M: YEASEN 10505ES. Plasmid input：200 ng/20 μL, 50°C, 40 mins. Selected citations: [1] Reversible phase separation of HSF1 is required for an acute transcriptional response during heat shock. Nat Cell Biol. 2022;24(3):340-352. (IF:28.824) [2] A Selective Small-Molecule c-Myc Degrader Potently Regresses Lethal c-Myc Overexpressing Tumors. Adv Sci (Weinh). 2022;9(8):e2104344. (IF:16.806) [3] Phosphatidic acid suppresses autophagy through competitive inhibition by binding GAPC (glyceraldehyde-3-phosphate dehydrogenase) and PGK (phosphoglycerate kinase) proteins [published online ahead of print, 2022 Mar 15]. Autophagy. 2022;1-15. (IF:16.016)
Features:
- Simple: Seamlessly assemble and clone up to six DNA fragments in a single reaction
- Flexible: Design guidelines allow assembly into any vector of your choice
- Efficient: Efficient for ligation of one to six fragments
Hieff™ Gold T4 DNA Ligase (5 U/µL) - 10300ES
Hieff™ Gold T4 DNA Ligase can catalyze the formation of phosphodiester bonds at the 5' phosphate end and 3' hydroxyl end of adjacent nucleic acids at blunt or sticky ends of dsDNA when ATP is used as a coenzyme, and can also catalyze RNA to link with ssDNA or ssRNA in double strands, but cannot catalyze linkages between fully single-stranded nucleotides.
This product is suitable for labeling the 3'-end of RNA, circularizing RNA and DNA oligonucleotides, cloning cDNA, etc. Concentration is 5 U/µL.
It is suitable for nucleic acid operations such as labeling RNA 3 '-end, looping RNA and DNA oligonucleotides, and cloning cDNA. Application included: Cloning (Blunt or sticky end joining); A linker or adaptor can be added to the flat-end DNA.
Figure 1: Ligation of Lambda DNA digest using Hieff Gold T4 DNA ligase
