Where does BamHI restriction enzyme cut?
(1995). BamHI binds at the recognition sequence 5′-GGATCC-3′, and cleaves these sequences just after the 5′-guanine on each strand. This cleavage results in sticky ends which are 4 bp long.
Does BamHI have star activity?
After 50-fold overdigestion with BamHI, >95 % of the DNA fragments can be ligated and recut with this enzyme. Star activity: Conditions of low ionic strength, high enzyme concentration, glycerol concentration >5 % or pH >8.0 may result in star activity.
Does bamh1 produce blunt ends?
Restriction enzymes can create fragments with sticky ends, as is the case with the enzyme BamHI, or blunt ends, as with HaeIII (Table 8.1).
How does Dpn1 Digest work?
Dpn1 only digest the methylated DNA, not your pcr product. I guess it will not work much efficiently in the pcr buffer. If you are amplifying your gene of interest from a plasmid, sufficient plasmid to get some colonies is available in your PCR mix. To get rid of the plasmid you may use DpnI digestion.
What type of ends do BamHI and EcoRI produce?
EcoRI – recognises the sequence 5’GAATTC’3 – sticky ends. BamHI – recognises the sequence 5’GGATCC’3 – sticky ends.
What is the difference between NdeI and BamHI?
NdeI requires 8bp to cut, thus it is not possible to conduct a double digest with this vector. BamHI is also a move active enzyme and comes in a more concentrated form. In pET11a and pET11b, the BamHI and NdeI sites are 31bp apart, separated by the T7 Tag.
Can I perform double digestion with NdeI and BamHI together?
Single digestion with either NdeI and BamHI showed almost complete digestion. However once i perform double digestion with both of them, the result is same as yours, which is incomplete digestion. You can try to do sequential digestion.
What buffers are available for restriction enzymes in 2021?
Beginning April 2021, we will be gradually transitioning to buffers containing Recombinant Albumin (rAlbumin) for restriction enzymes and some DNA modifying enzymes. All information on the website has been updated to reflect this change. Find more details at www.neb.com/BSA-free. BamHI has a High Fidelity version BamHI-HF ® ( NEB #R3136 ).
Where can I find more information about BSA-free NdeI?
Find more details at www.neb.com/BSA-free. An E. coli strain that carries the NdeI gene from Neisseria denitrificans (NRCC 31009). One unit is defined as the amount of enzyme required to digest 1 µg of λ DNA in 1 hour at 37°C in a total reaction volume of 50 µl. Not sensitive to CpG, dcm, or dam methylation.