Questions tagged [plasmids]
Often circular segments of DNA that can replicate outside of the chromosomes. Generally found in bacteria.
25 questions with no upvoted or accepted answers
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Are there any alternatives to the Epicentre product Plasmid Safe?
I need to remove any traces of linear DNA (both single and double stranded) from a ligation reaction while keeping circular DNA intact. Up to now, I have used Epicentre's Plasmid Safe to do the job. I ...
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Gibson assembly - primer design with A and T rich regions
I have question about Gibson assembly. I have done it several times and it always worked okay for us, but now I want to assemble together a fragment which has sequence like this: 5'...
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When doing minipreps, do "good" plasmids produce more DNA than "bad" plasmids?
I've been doing minipreps for close to 10 years using the standard qiagen kits and DH5a e. coli. I typically do 24 minipreps at a time and then identify the good plasmids by enzyme digestion, PCR, ...
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Can you Interrupt a promoter region by inserting another promoter region into it?
I currently have a plasmid (pFPV25.1) which contains a promoter region with multiple cloning sights in it. I would like to place a new promoter I have isolated, and I was wondering whether I could ...
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Why is there a size limit on inserts that a plasmid can accept?
Throughout my undergraduate education I have been taught that plasmids can't carry very large inserts, but I have never been told why. Any insight would be greatly appreciated.
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When purifying plasmid DNA after cloning, do you get endogenous plasmid in addition to your vector of interest?
Since bacteria naturally contain plasmids, when you do a cloning experiment and purify the plasmids at the end, do you get plasmid naturally present in that bacteria in addition to your vector? The ...
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Which plasmid can express T7 RNA polymerase in mammalian cells?
Does anyone know what plasmid could express T7 RNA polymerase in mammalian cells, I would like to transfect it with other plasmid containing Gene Of Interest under control of T7 promoter. Thanks a lot!...
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What is the origin of plasmids?
As I understand it, plasmids, like mitochondria, have their own genetic material and are capable of self-replication.
According to Wikipedia: Plasmids are considered replicons, units of DNA capable ...
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Plasmid harbouring subunit S of type I R/M system
We have an assembly of a genome of a Lactobacillus species. The genome also contains a 7.5kb plasmid.
Aside from the "usual plasmid genes" the plasmid contains:
- toxin/antitoxin system
- Subunit S ...
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Is it possible to both extract DNA sequences from plasmids and fuse them using primers for Gibson assembly?
Maybe I'm overcomplicating this, but I'm having some issues with a construct.
So, I have three plasmids containing a total of eight DNA sequences that I need to fuse into one linear sequence. I plan ...
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How to specify a promoter in de novo gene synthesis service?
I am trying to use a de novo gene synthesis service (Genscript) and one part confuses me:
Where do I pick the choice of promoter? Or is the promoter choice automatic based on my plasmid choice?
e.g....
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Protocol to transform _Staphylococcus aureus_
I'm working on transforming Staphylococcus aureus and I'm
totally lost on bypassing the RM systems. I have a plasmid already constructed, but I think I want to pass it through DC10B E. coli so that it ...
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What backbone to use for Helper dependent adenovirus (HDAd)
I have been trying to design a plasmid for a helper dependent adenovirus but while looking i wasn't able to see a backbone specifically for HDAds so I was wondering if a general adenovirus backbone ...
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Which is the best results for Plasmid dna extraction on gel electrophoresis?
I did plasmid dna extraction and uploaded on the gel.
Well 1: marker
Well 2: nicked,circular,and supercoiled bands
Well 3: linear,circular,and supercoiled bands
Well 4: circular and supercoiled bands.
...
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Problems reading my gel electrophoresis result
This is an image of my gel electrophoresis result.
I used the alkaline lysis method to extract E. coli plasmid and put it on gel electrophoresis. Based on what I had learned, I expected to see a band ...
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Migration of cut vs. non-cut plasmid on gel
As you might know there are 3 common forms of a plasmid:
ccc-form (CCC), being supercoiled.
oc-form (OC), being nicked and therefore relaxed.
linearized form (L), being cut on both strands and linear....
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Production method of custom seed mir451? How to edit middle of a fragment?
I am trying to create a plasmid that produces mir451 targeting a new gene, so I need to alter the seed sequence. The seed sequence is 7 bp long, so I think that is too long for PCR mutagenesis. How do ...
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Can the genetic sequences for CRISPR components be inserted into the host genome so that the cell perpetually produces CRISPR components?
Theoretically speaking, can you insert the gene sequences for cas9, sgRNA, and promoters into the host genome so that the cell perpetually produces CRISPR components?
In this scenario, I'm guessing ...
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Cloning in a large DNA fragment into a plasmid
I need to clone in a 30kb DNA fragment into my plasmid (~7kb).
Working with such a large fragment I have run into a few problems. The first was purifying it from the PCR mix used to amplify it out of ...
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Is there any commercial vector technology platform for boosting recombinant protein production?
I am a novice in biomedical industry. Our team would like to search new vector technology for boosting recombinant protein production. It's difficult to google a firm that provide those plasmids.
Dose ...
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How do we regulate the production of proteins when designing plasmids?
I think it should be no surprise that I, as many others, am interested in the new COVID-19 vaccines being developed. In my region of the world there are two mayor candidates. One is mRNA based and one ...
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Induce plasmid acquisition in bacteria to make them susceptible to antibiotics
Is it possible to use plasmids as a way of creating antibiotic susceptibility in bacteria.
For example; in Microbiology, an introduction 13th edition by Pearson , Page 230 "the researchers traced the ...
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Join a linear plasmid just using a primer (without ligase enzyme)
Can I use just a primer and PCR to join the cutted plasmid like this ? without using any further enzyme like ligase.
After the linear dsDNA plasmid annealed into 2 ssDNA, primer will bind to middle. ...
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History of plasmid maps
To create a small history of visual representations in biology, I am seeking the earliest published papers or books that have used plasmid maps as figures ; or strategies to effectively search for ...
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Can features of modified plasmids be divided into prokaryotic features and eukaryotic features?
Here's what I understand and please correct me if I am wrong:
Plasmids modified for gene therapy or genetic engineering should contain factors for certain functions in prokaryotic cells. For example, ...
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