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Questions tagged [plasmids]

Often circular segments of DNA that can replicate outside of the chromosomes. Generally found in bacteria.

14 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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57 views

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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146 views

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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304 views

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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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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35 views

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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99 views

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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21 views

How can I improve efficiency of Ecoli transformation?

I am an intern in biology institute. I have a 17.3kbp plasmid need to transform to Ecoli. But I have tried many time but have no or very few colony on LB plate. I know large plasmid have less ...
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35 views

Yeast integrating vectors: where to cut?

I understand that yeast integrating plasmids (YIps) must be linearized in order to promote homology-directed recombination. However I don't quite understand where. In the end, the external regions of ...
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Are there vectors available without any promoter?

Looking for a vector with the following characteristics: 1. Without any promoter 2. Selectable marker: Puromycin 3. Blunt end restriction sites 4. Transfection host: Human or Mouse cell lines.
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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 ...