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Restriction Digest Analysis 

Addgene
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In this video, we will digest a plasmid of known identity with restriction enzymes, run the products on an agarose gel, and analyze the results. We’ll focus on identifying the bands produced under different conditions and use the results to make conclusions about the plasmid.
Note: XbaI is effected by Dam methylation - see this blog post for more information blog.addgene.org/plasmids-101-...
For the full written Restriction Digest Analysis protocol, visit:
www.addgene.org/protocols/dia...
________________________________________________________________________________________________
0:00 Intro
1:00 Determining Expected Results - Control
3:21 Determining Expected Results - Digestions
4:31 Run the Gel
5:40 Comparing the Data

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30 июн 2024

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Комментарии : 44   
@addgene
@addgene Год назад
Addgene does not regularly monitor comments posted here, so we may not see your question immediately. We’d be happy to answer any questions sent to help@addgene.org as soon as possible. Please include the name of the video along with any questions so our support team can help. Thanks!
@romimoirangthem9277
@romimoirangthem9277 3 года назад
By far the best result analysis , explained it so easily
@catherineespinoza5917
@catherineespinoza5917 6 лет назад
This is very useful. I am sharing this with my undergraduate students to help them understand their lab results.
@addgene
@addgene 6 лет назад
That's great to hear Catherine! Please let us know if there are any other videos that you'd think would be particularly helpful.
@milokenneth6303
@milokenneth6303 2 года назад
You prolly dont care at all but does someone know a way to get back into an instagram account?? I was dumb forgot my password. I would appreciate any tips you can give me
@novelas3536
@novelas3536 Год назад
@@milokenneth6303 Bro what
@start5256
@start5256 5 лет назад
You people are doin wonderful job great really crystal clear
@kat-thee111
@kat-thee111 5 месяцев назад
This is such a clear video and so well explained. Very easy to follow, thank you
@hannahchen9272
@hannahchen9272 6 лет назад
Thanks for uploading this video, it helps a lot in interpreting the results I got from my first 200-level biochem lab, and would be more confident about what we are going to analyse in the following lab.
@addgene
@addgene 6 лет назад
Glad to hear you've found it useful Hannah!
@GoldbergDaBoss
@GoldbergDaBoss 4 года назад
This was great! You just saved my genetics lab report
@addgene
@addgene 4 года назад
glad to hear! we also have more detailed protocols over at addgene.org/protocols/
@nbent4607
@nbent4607 3 года назад
Direct to the point... I like it!!
@crys1537
@crys1537 7 лет назад
Really helped me out for my internship thanks alot !
@addgene
@addgene 6 лет назад
Glad to hear you found it useful!
@EatingWithEssence
@EatingWithEssence Год назад
Great explanation!
@rj6110
@rj6110 4 года назад
lots of helpful stuff here but im most impressed with the ability of the presenter to draw a perfect circle freehand lol
@hebaemam
@hebaemam 7 лет назад
very helpful video
@mohamedelmasry6388
@mohamedelmasry6388 4 года назад
you're awesome ♥️
@BoyMonkeyKid
@BoyMonkeyKid 3 года назад
laughing while learning should be illegal
@Hoxgene
@Hoxgene 3 года назад
this was amazing
@mahdis.the.curious
@mahdis.the.curious 8 месяцев назад
Having both nicked and supercoiled bands in lane 2 indicates that some of our plasmids stayed intact, and some others got nicked when undergoing the same conditions?
@raselbarua4578
@raselbarua4578 2 года назад
useful video, keep going
@anjaligupta6180
@anjaligupta6180 Год назад
For Age1 and xba1 double digestion (lane 5) How much nanogram of plasmid DNA was required to see the 300 bp band on gel. ?? If i have my estimate Dna conc around 25 ng/ ul post boil prep.
@smilykhajuria4411
@smilykhajuria4411 Год назад
Hy ..what is the procedure for ordering the plasmids from your site
@hebaemam
@hebaemam 7 лет назад
you are so funny guys
@bluebyrne7864
@bluebyrne7864 5 лет назад
Thank youu
@henkdevries2002
@henkdevries2002 3 года назад
What is the song in the intro?
@CestlaBios
@CestlaBios 5 лет назад
How can I know the concentration of restriction enzyme needed for this characterization?
@addgene
@addgene 5 лет назад
The concentration of the restriction enzyme will be on the tube of enzyme, usually in "U/mL" or "units/mL." Usually using ~0.2- 0.5 uL of enzyme will work. For more details see this protocol: www.addgene.org/protocols/restriction-digest/
@divyagautam4115
@divyagautam4115 2 года назад
If I have two bands showing in a single well during agarose gel electrophoresis, after RE digestion what does that mean
@addgene
@addgene 2 года назад
Thanks for the question. If you observe two bands on a gel after a restriction digest with a circular DNA molecule, such as a plasmid, that means your restriction enzyme(s) have cut two sites on your DNA molecule, producing two linear fragments. If you see two bands after a digest on a linear DNA molecule, that means your restriction enzyme(s) have cut only one site on your DNA molecule, producing two smaller linear fragments.
@srinathkshultz9968
@srinathkshultz9968 3 года назад
why only thosetwo enzymes ? why not gowith other restriction enzymes ki ecor1 or hind III
@addgene
@addgene 3 года назад
The enzymes shown in our video are for illustrative purposes, but within a plasmid sequence there are numerous restriction sites that you could consider using. On our plasmid webpages, unique restriction sites are annotated in bold on the plasmid maps. If you are performing a digest that is expected to return multiple bands, we recommend first confirming that these will be of sizes that can be easily separated on an agarose gel.
@mukhodenimatodzi3844
@mukhodenimatodzi3844 Год назад
Univen students 👩‍🔬 where are u?
@someguy1576
@someguy1576 5 лет назад
Why those enzymes in particular?
@addgene
@addgene 5 лет назад
Hi, these particular enzymes were just used as an example. It'll differ depending on the sequence of your plasmid which restriction enzymes you should use to cut the plasmid.
@srinathkshultz9968
@srinathkshultz9968 3 года назад
@@addgene what is the basis to choose those enzymes?
@guihuiqin6679
@guihuiqin6679 3 года назад
Cute~
@ericfeldsott7481
@ericfeldsott7481 2 года назад
he's wearing shorts in the lab-- rookie mistake
@zangetsu6638
@zangetsu6638 6 лет назад
4:55 #metoo that dude is gone!
@cinnamonbun216
@cinnamonbun216 4 года назад
Not funny
@zangetsu6638
@zangetsu6638 4 года назад
@@cinnamonbun216 and 5:11 too i think the dims have found biden's running mate
@raunakgupta2148
@raunakgupta2148 2 года назад
Oh well that was 2018 :, )
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