A friend and I were discussing the possibilities of doing some ball python genetic engineering to add a GFP or luciferase gene (harmless, but would make for some cool pets) and figured out the easiest way to do that would probably be to edit the sperm. Has anyone ever done artificial insemination with ball pythons? Could it be done? I know this is an unconventional question, but since we regularly do it with mice it seems like it might be possible and intriguing
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Yes! it may take me some time to dig up but I have definitely seen something along these lines.
As far as I can remember, the females egg ended up being the best place to start with rather than the sperm but I don’t want to misquote it so I won’t until I find the study.
I’ve discussed the use of CRISPR with @t_h_wyman on here before Genetics/morphs if you want to check that out.
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Interesting, thanks! I know there is a lab here that offered to sequence a ball python for me as they’re in the process of sequencing an entire zoo’s worth of animals just out of curiosity. However I’m not sure how useful that would be given that we’d have to sequence multiple normal/wild type snakes to generate a consensus sequence, then multiple of every morph in order to find the common SNP/mutation that causes the phenotype. It would be a wildly time consuming and expensive project.
Which is why I figured it might be interesting to instead try to add a gene that generates pigment or in this case, fluorescence. I was debating the use of CRISPR vs AAV infection to add the transgene, because while usually it is the egg that is edited, I thought the sperm might be easier access. I’m definitely going to have to look into this more
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Yeah and just to add to what you said, I have seen TONS of animals genetically engineered so I dont think it would be all that different from ball pythons. If I remember correctly a lab created a glowing anole lizard as part of a experiment
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I’m going to throw this here because it’s a really helpful search function that I don’t feel is well known.
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I feel like pubmed is better, google scholar pulls in all sorts of weird books and stuff. I asked on here after searching because I feel like the majority of people who are in ball python breeding aren’t publishing scientists and therefore there probably isn’t exactly a Nature article on it.
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Sometimes it’s them weird books that you find your answer 
I get what your saying though, a lot of the time it’s a struggle to find exactly what your looking for.
Well let me introduce you to my good friend @t_h_wyman 
Hopefully he will pop up and help us out.
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Yay, thank you! I saw some of his comments on another threads about ball python genetics and would love to get his opinion on genetic editing of snakes
We need a “genie from the lamp” emoji for when I get summoned LOL
@john can you make that happen??
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I discussed this topic with Dave a couple three months back so I will just plonk this here:
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And here is the paper we talk about. It is OpenAccess (i.e., no paywall) so go ahead and go crazy 
https://reader.elsevier.com/reader/sd/pii/S2211124719310058?token=C32438E1E63F0B9449F1FF0C60DF3364852DE294D90AE6C5A1F12C664E14763621DFF434339E78B1929E722E1489A86F
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This is exactly what I’m talking about, you seriously are priceless. I had to watch it in 3 parts because… kids 
Thank you.
This has genuinely been one of the most helpful things I’ve watched in regards to the morph Encyclopedia, I’m going to have to rewatch and take a lot of notes.
The way you describe the “ladder” of filters that pigmentation needs to climb is almost identical to a diagram I drew up a while ago that I was going to use as a example while asking you a question -
If we managed to map the genetic coding of a piebald, would it be possible to flip the lack of melanin and bombard it, so that we ended up with black patches instead of white?
As for CRISPR, I definitely see it being introduced to the hobby at some point in the near future.
I can’t see a arguement against it. Yes we may cause unwanted traits along the way, but that not only helps in further work, it will give us a understanding of how these same “edits” would react with other living things.
We already breed a whole bunch of morphs that have issues that we know about and have evidence for, maybe we can find a way to flick the “wobble” switch off/on.
Personally I think the reptile world will be full of a lot of crazy new things in 30 years time.
If you ever do any more of these talks, please give us all a heads up so we can tune in.
Here are a few links that may help simplify what you talk about at certain points.
The structure of crystals and how they affect the colour:
CRISPR:
(I know this one will hurt you to watch Travis as it’s very simplified, but it’s a good starting point for people that are interested)
Time to dive into the paper 
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Something else just popped in my head while reading. Would it be possible to manipulate a recessive mutations into being a dominant mutation?
For example a piebalds offspring needing only one copy from one parent to become a visual.
Also you mentioned that we could turn the"leg switch" for boas back on. If we did that would it be a instant thing, where the next clutch has legs, or would it take generations for them to return?
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Piebalds are not a melanin dysfunction mutation, they are a total pigment deposition dysfunction mutation so you would not be able to get that phenotype out of tweaking the gene responsible for Pied. Depending on which gene is responsible for BlkPastel or Mahogany, you might be able to do it with one of those (very very very heavy emphasis on MIGHT there)
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Doubtful. The wobble is intrinsic to the mutation so any correction that would cure the wobble would negate the phenotype associated as well
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I will try to remember. May have one coming up in the not too distant future but not 100% sure. But if you are interested you can find the couple episodes I did with Morelia Python Radio and there are another couple I did with From the Ground Up. And if you have some way of performing magic I also did a few with both Nick Mutton and Royally Addicted on the HerpNation Radio but I think those may be lost to the void. Likewise there was one episode of Reptile Radio I did, but it is also likely gone to the realm of electronic daemons
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Small clarification that how butterfly (and also bird plumage) crystal structure works is different than how the crystal lattices in reptiles and amphibians work. The broad overview is likely similar, I just do not want people to think it is an exact one-to-one comparison
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So long as it is not promoting conspiracy theory garbage I am okay with it LOL
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Broadly speaking, no. Recessive mutations are almost always loss-of-function type mutations. As such, a single allele’s expression being lost is compensated for by the WT allele continuing to be expressed.
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Yes and no. You would not get a boa that suddenly had a fully developed iguana sized hind legs because the evolutionary process that stunted the legs was the result of cumulative mutations that both shrank and “turned off” the developmental program. But with repeated edits to correct each of those mutation you could eventually get the full leg phenotype. Assuming no other problematic counter-expression, like what happens when you turn on tooth development in chickens… 
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Damn and I thought I knew alot about genetics, you and @t_h_wyman seem extremely knowledgeable as well. Glad we got guys like you on here to help fuel discussion
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While one day designer pets, including ball pythons, might be common, I still think we’re pretty far away. We don’t even have a sequence for the ball python genome, and in order to make edits using CRISPR, we would need to know exactly what genes to target. Even then, it’s not always as easy as it sounds. There’s a lot potential for off target effects. We’ve been making transgenic mice for years and still sometimes it doesn’t work, even for people who are very experienced. Just last year I tried to have a gene knocked out for study and not only did the knockout fail, but the mice ended up wildly obese and dying early, so evidently we affected something else.
Which is why I was thinking of starting with viral integration of a GFP transgene. It’s more imprecise and the random integration could cause unexpected effects based on where it lands, but would be an interesting place to start. It would also have the added benefit that if GFP integration caused an unexpected phenotype, we could sequence that region to make an association.
Is the spider wobble neurological or muscular, or a combination of both, or do we not know yet?
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Well, I mean it is kind of sort of my day job so… 
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There are a couple ball genomes out there. They have not been deposited in GeneBank, but they are out there, you just have to know the right guys. None of them are annotated though so… 
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CRISPR off-target effects are actually less frequent than the totally random viral integrations. Also, since we know the sequence for GFP, any off-target CRISPR integrations that generated an unexpected phenotype could be primer walked/sequenced just as easily.
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Definitively know? No. As you observed, no one is making genome-level interrogations of balls. But conventional wisdom is that we are dealing with a melanocyte migration dysfunction in the neural crest so odds are that it is a neurologic defect secondary phenotype associated with the base mutation(s) in the “wobble” morphs
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Haha yeah I almost forgot that you specialize in that field
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Mine as well, finishing up my doctorate in genetics at one of the better genetics programs in the country. Although I’m more neurogenetics than animal science genetics
The annotation is the hard part, getting the sequence is easy. A friend’s lab offered to do it for me, but I’m not really going to spend the time trying to sort through it.
That’s my point exactly, I’m ok with the random integrations because it would allow for sequencing of regions that generated a phenotype when disrupted. I could generate cool GFP+ pythons and start annotating if I wanted to. I already have both high titer AAV and lenti with GFP, so it seems like path of least resistance. I was merely pointing that we’re still a ways away from being able to cleanly generate morphs of our choosing via CRISPR
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I wouldnt say I’m extremely knowledgeable, Id say I just know enough to understand and ask what I don’t. I’m learning though.
Oh I completely agree, I don’t see us being able to say “make that snake purple” and we can just go ahead and produce a purple snake. Without investment from a billionaire I don’t see that becoming a reality until we can literally put a snake inside of a computer and break it down.
I do however see random test specimens becoming public. Such as what happened with the Glofish.
If you yourself did some mucking around and produced a completely brand new phenotype, you can expect all the big names in the hobby to be at your door with their cheque books… “Worlds first genetically modified ball python”.
What would someone benefit from not adding it to GeneBank? Why would they go to that effort and not make it useable?
Well played 
you will be a real asset to the whole community.
Again I completely agree. Also as you mentioned earlier or in a separate post, we would need multiple of each morph to create any sort of viable data.
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Eh, neither my formal education nor my career are animal oriented. Genetics is genetics is genetics. Sure, there may be species or family specific quirks but broad picture everything operates the same be it mouse or moose, tree or terrapin, slime mould or serpent.
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Depends on the researcher. Some keep hold of them because they still have work on the project and do not want others to scoop them. Others hold them back because they know the draft they have is kind of crummy and they would rather not have their name associated with a fragmented, shallow-depth draft. And some people are just lazy LOL
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