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Choosing Your Perfect Empty Backbone

Posted by Lianna Swanson on Aug 19, 2014 11:39:33 AM

Vectors (or empty backbones) are frequently used in molecular biology to isolate, multiply, or express the insert they carry in the target cell. These vectors allow you to test the function of Your Gene Of Interest (YGOI) in a controlled environment under various conditions. The first thing you'll need to decide when running your experiment, is which vector will best suit your needs?

At Addgene, we have a vast collection of empty backbones that have been designed, tested, and published by academic scientists. To help you find the vector that fits your experiments, I've described below some of the most frequently requested vectors in our repository and will discuss some of the features you may want to consider as you make your choice.

The first and most important thing you need to know is your expression system or environment. The host organism will determine the type of vector that you will need. You will also have to make sure that your plasmid has been incorporated into the host organism, usually achieved with the proper selection marker or antibiotic resistance.

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Topics: Plasmid How To, Plasmid Technology, Plasmid Elements

Synthetic Biology & the Importance of Assembly Standards

Posted by Guest Blogger on Jul 8, 2014 11:06:00 AM

The field of synthetic biology has seen tremendous growth in recent years. At Addgene, synthetic biology deposits have grown exponentially, from just 2 plasmids in 2005 to 439 plasmids deposited last year. To shed some light on this growing field, we asked our friends at iGEM to share their expertise and discuss the importance of standards in the field.

The following post was contributed by Kim de Mora, iGEM Fellow.

What is Synthetic Biology?

“What I cannot create, I do not understand.” – Richard Feynman

This Feynman quote perfectly embodies the aims of synthetic biology in a single sentence. During the history of humanity, some of the most complex devices we have constructed are nuclear submarines, the space shuttle, the international space station and the Internet. But in all our existence, we have yet to design, engineer and build a cell from the ground up. A single bacterial cell is orders of magnitude more complex that the aforementioned feats of mechanical, aeronautical, electrical and computer engineering. These devices could be built because the underlying physical model of how the world works is understood by scientists and applied by engineers to practical ends.

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Topics: Plasmid Technology, Synthetic Biology

Plasmids 101: Luciferase

Posted by Jason Niehaus on Jun 24, 2014 11:59:00 AM

Luciferases are a class of enzymes capable of catalyzing chemical reactions in living organisms resulting in the emission of photons. The most familiar bioluminescent organism for most people is the firefly (Photinus pyralis) and perhaps not surprisingly it is also the most commonly used bioluminescent reporter. This beetle emits a yellow-green light with a peak emission at 560nm. Shortly after the initial article describing the cloning of firefly luciferase was published in 1985, several studies utilized luciferase as a genetic reporter in plant and mammalian cells. Luciferase assays have since become a gold standard in gene expression analysis and a luciferase gene (one of many available to choose from) is now a common feature in reporter plasmids. 

Learn How Luciferase Can Be Used In Concert with Fluorecent Proteins in Nano Lanterns

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Topics: Plasmid Technology, Plasmids 101

Designer PUF Proteins for Any RNA Target

Posted by Kendall Morgan on Jun 17, 2014 3:57:00 PM

With the meteoric rise of CRISPR technology, the ability to direct enzymes – from nucleases to transcription factors – to specific sequences of DNA has become commonplace. This ability has opened up a world of possibilities in the engineering of complex gene networks. A comparable system for targeting specific sequences of RNA is highly desirable for extending the complexity of genetic circuits, allowing for tighter spatio-temporal control of gene expression within a cell. Thanks to the work of Huimin Zhao and colleagues, we now have just the tool…designer PUF proteins!

A newly available PUF Assembly Kit makes it possible to devise RNA binding proteins to hit any target of interest. The new tool was developed and implemented by applying the Golden Gate cloning method to human proteins known as Pumilio/fem-3 mRNA binding factors (PUF). In a single step, researchers can now assemble designer PUF domains for RNA specificity engineering.

“The RNA binding domain is interesting because by changing certain amino acids you can change the specificity,” explained Zhanar Abil of the University of Illinois at Urbana-Champaign.

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Topics: Plasmid Technology, Synthetic Biology, Plasmid Kits

Interview: Nicola Patron on Plant Synthetic Biology, MoClo, and More

Posted by Kendall Morgan on Jun 12, 2014 11:30:00 AM

Nicola Patron is Head of Synthetic Biology at the Sainsbury Laboratory, where she often feels more like an engineer than a biologist. Their focus at the lab is on plant-pathogen interactions, and her aim is to produce constructs and edit genomes so as to make plants, and agricultural crops in particular, resistant to disease. They also devise biosensors designed to elucidate the molecular interactions that go on between plants and their pathogens.

As Patron explains it, her work has always been focused on gene transfer, from transgenes to plants, chloroplast to the nucleus, or pathogens to their hosts. I spoke with her about what motivates her research, the MoClo Kit she and Sylvestre Marillonnet share with the scientific community via Addgene, the struggles of plant scientists and how they work to overcome them, and why she spends some of her time engaging with others on Twitter, among other things.

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Topics: Plasmid Technology, Scientific Sharing, Synthetic Biology, Plant Biology, Plasmid Kits

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