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Questions tagged [cloning]

The process in nature or in the lab by which a new organism is created that is genetically identical to its predecessor.

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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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Ligation without purifying insert

I am planning to insert a 45 bp sequence in a vector. After restricting my insert to create compatible sticky ends, I am finding no way to clean it up. Is there any way for cleaning this fragment ...
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How can seedless fruits not be GMO?

Biology is the closest I could find to botany on SE. Grafting can not occur naturally and so if a fruit does not have seeds there is no way it could reproduce in nature. That logic makes complete ...
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Why is DNA polymerase added at the end of PCR reaction?

PCR reaction is used to amplify DNA fragment. Each reaction requires a DNA template, buffer, dNTP mix and a unique pair of primers, one ''forward'' primer and one ''reverse'' primer. The DNA ...
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use of antibiotics in selection of good plasmids

I have not understood the role of antibiotic medium in filtering the bacteria containing the recombinant plasmids with the ones having normal plasmids. Since all the plasmids of a certain bacterial ...
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Are clones less likely to be fertile?

I know that hybrid animals like mules are infertile. What about clones? Do cloned animals show the same fertility as their original?
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Insertion Confirmation Cloning Strategy

So I am in a bit of a time constraint. Essentially, I inserted a DNA fragment via molecular cloning which contains a unique RE site. I need to confirm whether my colony has or does not have the ...
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Thousands of Reporters Integrated in Parallel (TRIP) barcodes

Has anyone worked with constructing barcodes for TRIP? How does constructing of barcodes work in general? I don't seem to be able to grasp the basics of constructing these barcodes.
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Misuse of DNA leaks to create your clones, or children

Recently, sequenced DNA data leaks have been reported. The linked article discusses possible risks like blackmailing based on health related data. However there could be one more misuse, at least ...
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Are cloned spieces significantly more vulnereble to deseases than sexually reproducing species?

I would like to be able to compare the risk for species to go extinct implied by their reproduction mechanism in the very short term. Imagine we choose some species A that can reproduce both sexually ...
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Is it Theoretically Possible to Create a Male clone from the donor Cell of a Female?

I understand that Dolly the Sheep was created without the presence of a sperm cell. Instead, nucleus DNA from somatic cells of adult sheep were used. Since Dolly was female, i am assuming that the ...
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55 views

Which comes first, PCR or Gel electrophoresis?

If I want to find out wither a group of patients have the abnormal BCR-ABL cancer gene, how do I benefit from the PCR and the gel electrophoresis techniques? I'm kinda lost trying to determine which ...
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Polysporin/antibioltic ointment and agar plates

Has anyone tried applying a layer of antibiotic ointment to the surface of their agar after autoclave sterilization? A friend suggested that I use an antibiotic in my agar. So it got me wondering if ...
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Micro-propagation of Moss in Agar (sterilization?)

I'm having trouble sterilizing my moss sample for micro-propagation. Moss is basically a sponge and it has a massive surface area. I tried Isopropyl alcohol figuring it had a very high probability of ...
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Are drones from one honey bee queen all clones?

The Honey Bee queen lays eggs that develop into worker bees (female) if she fertilizes the egg. If she does not fertilize the egg, it will develop into a drone (male). The drone only has the queen'...
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Why is easy to clone cats but it's hard to clone dogs?

According to this article, Here is why we're not cloning humans there are species who are easy to clone and other who are hard to clone. Cats and mices would be in the easy to clone set , and dogs and ...
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Isolating gene of interest from a large strand of DNA [closed]

I was always confused about this in class and never learned it fully. So say you have 10,000 bp of DNA and you just want to isolate 100bp of specific region(gene of interest) using restriction enzyme. ...
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PCR gene cloning

I've been set some homework and I have no idea where I would start. Any help would be appreciated. Thanks Which version of primer 1 should be used to allow the maf gene to be cloned into the pQE-30Xa ...
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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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Cloning problem. Digestion test at the end of the cloning is always negative

I have a problem with my cloning, I'm trying to clone a 483 pb in a pSecTag vector with restriction sites HindIII and BamHI. Here are the procedures I'm doing: • I introduce the restriction sites ...
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Somatic cell cloning always produces female offspring?

I was looking at the somatic cell cloning of goats. One of the advantages of the process said was All offspring produced are female. Now I'm confused. How is this the case? I know that the ...
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Cloning DNA fragment - at least trying to

I`ve been trying to clone a fragment (about 1.6 kb) using pGEM system, unfortunately it was not succesful. Then, I moved to pJET, and guess what? Again, very low transformation efficiency. Now I am ...
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If a woman was fertilized with her own DNA, would it be her clone?

Several questions: 1) Can human DNA fertilize an egg, without having been naturally packaged in a sperm cell? 2) If a female had her ovum fertilized with her own DNA, would the fetus be her clone? ...
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Plasmid transformation yields only empty vectors

I have to electroporate pMAD cloned with an insert of 7.4 kb in my S. aureus isolate. When I electroporated in RN4220 I had no problems but when I put it in my isolate I just recover few colonies and ...
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LacZ' selection: blue colonies despite ligation of insert

It has been suggested that bacteria transformed with pBlueScript vector, containing an insert in the middle of the lacZ' gene, can still give blue colour on X-gal, if the insert is small and ligated ...
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What step of the cloning process requires an egg cell? Have alternatives been attempted?

In the famous Dolly the sheep an egg cell was required. Why was specifically an egg cell required compared to say sperm cells or any other cell types? Given that egg cells are not trivially harvested ...
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Genetic information storage and the extent to which it controls appearance/structure

So there are often articles about cloning from a small piece of tissue etc. My question is (two actually) Is the Complete information about entire organism's structure stored (as in duplicated) all ...
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178 views

Complementation cloning: what, why and how? [closed]

I get that you're cloning a whole cDNA library, but why does E. coli need a complementary gene to allow the gene expression? Why is that complementary gene considered to be a mutant? AND how does ...
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Can a human be made with ovum but without sperm?

This article says that scientists were successful in making a mouse using only a sperm and no egg (ovum). The article also states that this procedure could be applicable to humans. However, I want to ...
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1answer
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Difference between clonal and subclonal mutations

I'm a physicist writing a proposal that has to do with cancer as a disease driven by evolutionary selection. As far as I understand, all tumors start with a single precursor (single cell or group of ...
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47 views

Why doesn’t p53 cause the repair of cellular DNA that has been altered experimentally?

When scientists mutate bacteria or even embryos of lambs and other animals, why doesn't the p53 reverse whatever mutation the scientists cause? I know that the p53 stops the DNA from mutating and ...
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2answers
572 views

When is strict sterile technique necessary? Cloning vs. protein expression

Background: I have previously worked with RNA, and then we use laminar flow hoods for any work where we do plating or inoculation of cultures while applying sterile technique. I am now working in a ...
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1answer
3k views

Maximum loading volume for agarose gel with 10 well comb

I want to load my agarose gel with my PCR products, which are 50 µL reactions. I am wondering how much of those reaction I can load into one well if I use a standard agarose mini gel (10 well comb, ~...
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Can you make sperm and eggs from the same person using in vitro gametogenesis?

In a recent popular science article there is a discussion of the potential disruptive technologies that could become available with in vitro gametogenesis (IVG). Using skin cells you will be able to ...
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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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Is a fully functional hermaphrodite possible in humans? [closed]

Is it possible for some one to achieve asexual reproduction if they had complete fully functional reproductive system? What problems might they encounter during conception and fetal development? If ...
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1answer
432 views

Is a Linker sequence mandatory in primers for PCR Cloning?

I have designed the primers for PCR cloning, but i was not getting any colonies after the ligation. One of our colleagues said, that the restriction digestion might not be working as there is no ...
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What is the typical amount of full length clones in a cDNA library?

For a Yeast Two Hybrid project we need to make a cDNA library from a single celled organism. We used the Cloneminer II kit from Thermo. We followed the manual and performed all the quality control ...
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Why we need an egg cell donor and a surrogate mother in an organism cloning process?

Could a whole set of artificial organism-cloning process only be operated on the somatic nuclear donor ,which means for example ,can we extract the somatic nuclear from animal "A" and put it into the ...
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Why does the cloned cat, CC, exhibit mosaic fur color?

Since fur color in this cat is from a sex-dependent gene, I see how calico cats occur. However, I don't fully understand why it happens in cloned cats. When the cat is cloned, the DNA is taken from a ...
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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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Methods for varying insert size in E. coli clones

I'm writing my master's thesis proposal, and I need to improve the solution-behavior of some truncated protein domains which have previously shown poor solubility when expressed in E. coli. To ...
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2answers
3k views

Does a woman contain all the genes needed to make a man?

I know the answer of inverse is at here, but how about this question? I also read this question, can I imply to human that a woman can also contain all the genes needed to make a man? Edit: I think ...
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2answers
9k views

Are identical twins clones? [closed]

Identical twins have the same genetic make up but they differ in development so are they in a way clones?
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57 views

What's the rationale behind freezing one's eggs if cells can be cloned?

It seems to me there's a lot of discussions around the topic of somebody freezing her eggs. What I don't understand though are the following: Why would you freeze your eggs when you know that the ...
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UV light-induced mutagenesis during gel extraction

this is a very short question I did not find the answer for online, neither on this nor other fora. At the beginning of my cloning protocol, I extracted the band with the sequence of interest directly ...
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1answer
106 views

Using PGAL10 promoter to overexpress specific yeast genes: How to?

I was reading a poster about expression boosting for Artemesinin production in yeast and it had this line: PGAL1 and PGAL10 have been used for the overexpression of all genes from the ...
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de novo gene synthesis vs cDNA library creation from plant RNA: Pros & Cons

What are the relative pros & cons of the following two approaches when one wants to insert a known, sequenced plant gene into a target host organism: de novo gene synthesis vs plant matter ...
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1answer
231 views

What are the problems in using an ssDNA or an mRNA for expression instead of using a plasmid?

Let's say you want to do a gene cloning experiment. You have an empty expression vector (mRNA or DNA) and are looking to clone some gene X into it. Why is it you can only buy X already inside a vector ...
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Unwanted E. coli product in plasmid product

Using a TOPO XL Cloning kit, our lab took some amplified human DNA, inserted it into the vector, and attempted to clone it in E. coli. However, upon sequencing the purified plasmid product, the ...