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  • Any newcomer who joins a molecular biology lab will undoubtedly be asked to

  • design...

  • construct...

  • or modify a plasmid.

  • But what exactly is a plasmid, and why is it so useful in the lab?

  • At their most basic level, plasmids are small circular pieces

  • of DNA that replicate independently from the host from a seminal DNA compared to

  • the millions or billions of bases that make up the entire genome plasmids

  • typically contain a couple thousand at most.

  • They're relatively small, stable,

  • and easy to manipulate

  • So where do plasmids come from?

  • In nature they're found in microbes, like bacteria.

  • In the 1940s scientists, began noticing

  • that there are heritable cytoplasmic factors that could be transferred between cells.

  • But there was no consensus on what they were or what to call them

  • It wasn't until 1952 that Nobel laureate Joshua Lederberg coined the term plasmid

  • a combination of the words cytoplasm and id (Latin for "it").

  • In nature plasmids often contain genes that provide a competitive advantage,

  • giving its host bacterium an ability that it didn't have before

  • These benefits include antibiotic resistance,

  • the wherewithal to survive in harsh environments,

  • and even the ability to wage war to gain an environmental advantage.

  • But plasmids aren't just useful to bacteria.

  • because they're incredibly easy to manipulate,

  • they've also made themselves indispensable to

  • life scientists and bioengineers.

  • Plasmids created in the lab are known as

  • constructs or vectors.

  • To understand what plasmids can do let's break down its

  • parts using a plasmid map.

  • All plasmids contain an origin of replication, or Ori.

  • It tells the plasmid where to begin replication.

  • Plasmids often contain genes that are advantageous for survival.

  • One of the most common naturally occurring,

  • types of genes is antibiotic resistance.

  • When used in the lab, antibiotic

  • resistance genes allow scientists to separate out cells that contain plasmids

  • from those that don't.

  • The wonderful thing about plasmids is that they can be

  • easily engineered and can introduce foreign DNA into cells through electroporation,

  • or other methods.

  • Many plasmids are designed so scientists can

  • insert genes they want to be expressed in organisms.

  • One way to do this is through restriction sites.

  • Restriction enzymes recognize these sites and cut

  • out the present gene like molecular scissors.

  • Then a different gene can be inserted into the site.

  • Often these restriction sites are located in what's

  • called a multiple cloning site,

  • a short segment of DNA that contains several restriction sites.

  • This adds flexibility to the cloning process.

  • How is the inserted gene expressed?

  • A promoter site which is upstream of the inserted gene on the plasmid,

  • acts as a green light that allows gene transcription

  • RNA polymerase binds to the promoter moving along the strand.

  • As it moves along the strand, it creates a new strand of mRNA, expressing the gene.

  • The plasmid cloning process is very versatile so there's a whole array of genes that

  • scientists can introduce into the cell

  • for example if you wanted to track a specific species of bacteria in a

  • population you could insert a GFP expressing gene into a plasmid and

  • transform it into your bacterium.

  • Your cells will fluoresce making, them easier to find.

  • Or if you're trying to study the effect of a specific protein on a phenotype,

  • you could insert the gene of interest into the plasmid.

  • Move this into a cell,

  • and look for changes in the cell

  • We hope you enjoyed this video in our Plasmids 101 series.

  • To catch our newest videos, subscribe to the Addgene YouTube channel,

  • and visit blog.addgene.org for more plasmid info

Any newcomer who joins a molecular biology lab will undoubtedly be asked to

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