Long ssDNA Preparation Kit for 3kb, Academic
Product#: FNK-DS625-A
$956.80
Availability:
Ships in 1-2 Weeks
Long ssDNA Preparation Kit for 3kb
A simple and highly effective method for preparing a long single stranded DNA for CRISPR workflows.
Knock-in efficiency becomes dramatically better when CRISPR/Cas9 is performed with long single-stranded DNA
The long ssDNA prepared by our kits has a defined sequence and length with high fidelity enzymes and does not include mutation and terminal deletion caused by PCR.
Knock-in efficiency becomes dramatically better when CRISPR/Cas9 is performed with long single-stranded DNA
The long ssDNA prepared by our kits has a defined sequence and length with high fidelity enzymes and does not include mutation and terminal deletion caused by PCR.
Cat. No. FNK-DS625-A
Store RT,-20°C
Size 1 kit
Product ID and Size
| Cat No. | Size | Link |
| FNK-DS625-A | 1 kit |  |
| FNK-DS625-C | 1 kit | |
Description
The Long ssDNA(Long Single Strand DNA) Preparation Kits (LsODN Preparation Kits) provide a simple and easy method for generation of a long ssDNA (within 1,500 base or within 3,000 base). A long ssDNA prepared by this kit has defined sequence and length as it does not include inside mutation and terminal deletion caused by PCR, exonuclease side reaction, not high-fidelity reverse transcriptase reaction or not high-fidelity synthetic oligonucleotides.
The procedure is almost same as the method to obtain dsDNA fragments. The DNA of interest is cloned into a plasmid. The resulting plasmid harboring the DNA is digested with a pair of two nicking endonucleases or a combination of a nicking endonuclease and a restriction enzyme. The nicked plasmid is denatured by mixing with Denaturing Gel-Loading Buffer and then subjected to agarose gel electrophoresis. The band corresponding to a long ssDNA is excised and extracted with our kit (#DS640) specially designed for this kit.
The procedure is almost same as the method to obtain dsDNA fragments. The DNA of interest is cloned into a plasmid. The resulting plasmid harboring the DNA is digested with a pair of two nicking endonucleases or a combination of a nicking endonuclease and a restriction enzyme. The nicked plasmid is denatured by mixing with Denaturing Gel-Loading Buffer and then subjected to agarose gel electrophoresis. The band corresponding to a long ssDNA is excised and extracted with our kit (#DS640) specially designed for this kit.
Kit Contents
- DS615 Long ssDNA Preparation Kit for 1.5 kb (LsODN Preparation Kit)
- pLSODN-1 10 µg (0.5 µg/µl)
- pLSODN-2D 10 µg (0.5 µg/µl)
- Denaturing Gel-Loading Buffer 1 ml (100 loadings)
- Long ssDNA Gel Extraction Kit for 3kb (DS640) (25preps)
- DS625 Long ssDNA Preparation Kit for 3.0 kb (LsODN Preparation Kit)
- pLSODN-3 10 µg (0.5 µg/µl)
- pLSODN-4D 10 µg (0.5 µg/µl)
- Denaturing Gel-Loading Buffer 1 ml (100 loadings)
- Long ssDNA Gel Extraction Kit for 3kb (DS640) (25preps)
- A long ssDNA (within 1,500 base or within 3,000 base) can be prepared.
- A long ssDNA has defined sequence and length.
- Simple principle and easy procedure.
- High yield and high quality.
- The DNA of interest is cloned into a plasmid using a pair of two nicking endonuclease sites or a combination of a nicking endonuclease site and a restriction enzyme site.
- The resulting plasmid harboring the DNA is digested with a pair of two nicking endonucleases or a combination of a nicking endonuclease and a restriction enzyme.
- The nicked plasmid is denatured and then subjected to agarose gel electrophoresis.
- The band corresponding to a long ssDNA is excised and extracted.
Plasmid Map
Data
Fig. Long ssDNAs prepared by this kit
A 1.5 kb DNA fragment of interest was cloned between the Nt.BspQI and the Nb.BsrDI sites of pLSODN-1. Similarly, a 3.0 kb DNA fragment of interest was cloned between the Nt.BspQI and the Nb.BsrDI sites of pLSODN-3. The resulting plasmids were digested with Nt.BspQI and Nb.BsrD. The double nicked plasmid was mixed with Denaturing Gel-loading Buffer and heated, then loaded to conventional non-denaturing agarose gel electrophoresis. The band corresponding to a long ssDNA was excised and extracted.
Lane 1: The double nicked pLSODN-1 harboring 1.5 kbp DNA fragment
Lane 2: Purified long ssDNA (1.5 kb)
Lane 3: The double nicked pLSODN-3 harboring 3.0 kbp DNA fragment
Lane 4: Purified long ssDNA (3.0 kb)
A 1.5 kb DNA fragment of interest was cloned between the Nt.BspQI and the Nb.BsrDI sites of pLSODN-1. Similarly, a 3.0 kb DNA fragment of interest was cloned between the Nt.BspQI and the Nb.BsrDI sites of pLSODN-3. The resulting plasmids were digested with Nt.BspQI and Nb.BsrD. The double nicked plasmid was mixed with Denaturing Gel-loading Buffer and heated, then loaded to conventional non-denaturing agarose gel electrophoresis. The band corresponding to a long ssDNA was excised and extracted.
Lane 1: The double nicked pLSODN-1 harboring 1.5 kbp DNA fragment
Lane 2: Purified long ssDNA (1.5 kb)
Lane 3: The double nicked pLSODN-3 harboring 3.0 kbp DNA fragment
Lane 4: Purified long ssDNA (3.0 kb)
Reference
- Yoshimi K. et al., ssODN-mediated knock-in with CRISPR-Cas for large genomic regions in zygotes. Nat Commun. 2016 Jan 20;7:10431
- Wang Y.et al., Long noncoding RNA lncHand2 promotes liver repopulation via c-Met signaling. J Hepatol. 2018 Oct;69(4):861-872.
- Zhu P. et al., LncGata6 maintains stemness of intestinal stem cells and promotes intestinal tumorigenesis. Nat Cell Biol. 2018 Oct;20(10):1134-1144.
- Zhu P. et al., IL-13 secreted by ILC2s promotes the self-renewal of intestinal stem cells through circular RNA circPan3. Nat Immunol. 2019 Feb;20(2):183-194.
- Nozaki S, and Niki H. Exonuclease III (XthA) enforces in vivo DNA cloning of Escherichia coli to create cohesive ends. J Bacteriol. 2018 Dec 10.
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