Cre recombinase Puns

I'm reading a paper [(Link)] (https://onlinelibrary.wiley.com/doi/full/10.1111/gbb.12202) and I know the knockout is for an oxycontin receptor, and 5-HT is serotonin, but what does heterozygous Cre recombinase‐expressing; KO, 5‐HT Oxtr KO mean? I study neuroscience, not biology, in college, so explain like I've taken intro to biology if you can please.

I was wondering what the purpose of cre recombinase is in transgenic mice. I know what/how Cre recombinase works; however, I am a little confused on why it is used. A study I am looking at uses mice with different genes (vGAT-cre, vGlut2-cre, TH-cre, etc.) and subsequent injection of AAV-ChR2 (channelrhodopsin). I am confused on why Cre recombinase is used and why it is necessary. My preliminary assumption is that the Cre recombinase somehow aids in the delivery of the viral vector with channelrhodopsin, but I am not certain. Any explanations/article suggestion are much appreciated.

Thanks.

As a protein to cells, as a plasmid in cells, as mRNA in cells, etc?

When I did my PhD I did a lot of plasmid generation etc in areas I'd probably call "synthetic biology" if I were doing it now. >>10 kb constructs with repetitive sequence in recombinase deficient competent cells, Cre-lox inversion based expression control etc, all with REs and MCSs etc plus a bit of PCR if I wanted to insert loxP sites etc. We had just switched over from EtBr to Sybr Green, diagnostic digests in individual colonies, that sort of thing.

We're getting back into that area, and I want to make sure I'm teaching people to use current approaches rather than the old stuff just because I did it that way. I'll lay out a rough workflow below that captures most of what I've come up with (numbered to facilitate linking comments to individual points), please let me know any suggestions, if there's something even more up-to-date I should consider etc.

I'd like to keep costs down as much as reasonably possible so if an undergrad wants to try an idea for something I can say "go ahead, we'll see if it works".

1. Search AddGene et al before making anything
2. Verify important bits with sequencing
3. Whole-construct DNA synthesis where justified by price vs time / complexity of DIY
4. Semi-synthesis by inserting sequence into the 5' end of PCR primers and amplifying the whole plasmid then ligating to circularize (I used to do this up to ~120 bases into 16 kb plasmids, but I'm guessing more is no problem these days?)
5. Assembly of multi-component constructs via NEBuilder HiFi system (looked at a bunch of options but this seems like a good one to standardize on?) (do you need to gel-purify or can you just go straight from the PCR tube?)
6. in order of selection speed in mammalian cells puro > hyg > G418, avoid zeo due to risk of DNA damage / mutation - any really good new selectable markers these days?
7. Routine QC by slicing up the construct with a couple of 4-cutters so you get a lot of bands (I used to use MboI and RsaI in separate single digestions, sometimes TaqI for the thermostability)
8. run out on a 1-1.5% agarose gel with thin wells for sharp banding, stain with Sybr Green (is there something better now that's not a gimmick? if there's something you can use and visualize on the open bench without eye protection that'd be great e.g. neither toxic like EtBr nor requiring UV excitation)
9. are there any good/cheap alternatives to diagnostic gels? I used to play around with capillaries for a while for sequencing, there's got to be some

Рис. 1. Схема работы искусственной системы, имитирующей репликацию и эволюцию в живых клетках. А — общая схема эксперимента. Б — схема репликации и рекомбинации искусственного «генома» в сопряжении с транскрипцией и трансляцией на уровне индивидуальных молекул, обеспечивающими реализацию самопроизвольной эволюции последовательности ДНК в ряду «поколений». Подробности в тексте. Рисунок из обсуждаемой статьи в ACS Synthetic Biology

Японским ученым удалось создать предельно простую систему, воспроизводящую некоторые базовые свойства клеточной жизни — способность к самовоспроизводству и эволюции на основе дарвиновского механизма отбора. В этой системе присутствует искусственный «геном» (кольцевая ДНК), который размножается посредством репликации и рекомбинации. «Геном» содержит гены, которые кодируют ферменты, обеспечивающие эти два процесса (ДНК полимеразу фага phi29 и рекомбиназу Cre). Транскрипция и трансляция этих генов осуществляется готовым коктейлем ферментов и низкомолекулярных соединений, добавляемым в систему в каждом раунде репликации. Реакции протекают в искусственно формируемых микрокомпартментах — капельках водно-масляной эмульсии, что обеспечивает необходимое условие для конкуренции и адаптивной эволюции «генома». Мутации, накопившиеся в кодирующих и некодирующих участках «генома» за 30 раундов (поколений) самовоспроизводства, повысили эффективность репликации в системе за счет нескольких установленных в ходе исследования механизмов.

Создание искусственной системы, воспроизводящую свойства живой клетки, — одна из амбициозных задач на стыке химии и биологии, которые ставит перед собой современная наука. Эта задача представляет интерес в первую очередь в контексте стремления понять процесс возникновения первых живых клеток из неживой материи. Ведь, как говорил Ричард Фейнман «чего не могу воссоздать, того не понимаю». Во вторую очередь важен потенциал практического использования (а как же!) таких искусственных систем в качестве фабрик по эффективной продукции чего-нибудь полезного человечеству.

Обсуждаемая ниже работа была выполнена в лаборатории Токийского университета, которой руководит Норикадзу Итихаси (Norikazu Ichihashi). Лаборатория специализируется именно на создании искусственных систем, способных к самовоспроизводству и эволюции — то есть систем со

I don't want to step on anybody's toes here, but the amount of non-dad jokes here in this subreddit really annoys me. First of all, dad jokes CAN be NSFW, it clearly says so in the sub rules. Secondly, it doesn't automatically make it a dad joke if it's from a conversation between you and your child. Most importantly, the jokes that your CHILDREN tell YOU are not dad jokes. The point of a dad joke is that it's so cheesy only a dad who's trying to be funny would make such a joke. That's it. They are stupid plays on words, lame puns and so on. There has to be a clever pun or wordplay for it to be considered a dad joke.

Again, to all the fellow dads, I apologise if I'm sounding too harsh. But I just needed to get it off my chest.

Do your worst!

They were cooked in Greece.

I'm surprised it hasn't decade.

For context I'm a Refuse Driver (Garbage man) & today I was on food waste. After I'd tipped I was checking the wagon for any defects when I spotted a lone pea balanced on the lifts.

I said "hey look, an escaPEA"

No one near me but it didn't half make me laugh for a good hour or so!

Edit: I can't believe how much this has blown up. Thank you everyone I've had a blast reading through the replies 😂

It really does, I swear!

Because she wanted to see the task manager.

Heard they've been doing some shady business.

but then I remembered it was ground this morning.

Edit: Thank you guys for the awards, they're much nicer than the cardboard sleeve I've been using and reassures me that my jokes aren't stale

Edit 2: I have already been made aware that Men In Black 3 has told a version of this joke before. If the joke is not new to you, please enjoy any of the single origin puns in the comments

They’re on standbi

BamBOO!

A play on words.

Pilot on me!!

Christopher Walken

Nothing, he was gladiator.

Or would that be too forward thinking?

Dad jokes are supposed to be jokes you can tell a kid and they will understand it and find it funny.

This sub is mostly just NSFW puns now.

If it needs a NSFW tag it's not a dad joke. There should just be a NSFW puns subreddit for that.

Edit* I'm not replying any longer and turning off notifications but to all those that say "no one cares", there sure are a lot of you arguing about it. Maybe I'm wrong but you people don't need to be rude about it. If you really don't care, don't comment.

What did 0 say to 8 ?

" Nice Belt "

So What did 3 say to 8 ?

" Hey, you two stop making out "

When I got home, they were still there.

I won't be doing that today!

This morning, my 4 year old daughter.

Daughter: I'm hungry

Me: nerves building, smile widening

Me: Hi hungry, I'm dad.

She had no idea what was going on but I finally did it.

Thank you all for listening.

Where ever you left it 🤷‍♀️🤭