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Speech

Assalamualaikum and good morning to all. My name is Aminul Islam Apurbo, ID:________

First of all, I would like to thank our honorable faculty _______ for giving me this opportunity to present something to you.

Now, the topic of this presentation is about SHUTTLE VECTOR.

There is the contents that we are going to cover. First of all we will know about shuttle vectors, then we will discuss about the features or characteristics and the structure of a shuttle vector. After that we will discuss about how we can construct a shuttle vector and lastly the advantages and disadvantages of a shuttle vector. We will cover the first two topics in one slide, 2nd slide will contain the construction and in the 3^rd slide we will see about the advantages and disadvantages. Soh, this is our proceeding plan.

Now, let’s get started.

Before going to the definition, lets understand How Did The Idea of A Shuttle Vector Come.

Idea behind Shuttle Vector

We all know about vectors, right? Vectors are used as a vehicle to transfer foreign genetic material into another cell. We have known about many vectors already. Some vectors can propagate only in bacterial host cells and some vectors propagate in yeast or eukaryotic host cell. Will it be prokaryotic host cell or eukaryotic host cell, that is very specific, right?

Now, imagine that you have a vector that can propagate in both organism whether it is prokaryote or eukaryote, the vector can be manipulated in both host cells. We can use a single vector for bacteria as well as yeast. It will be much more appreciating, right? That’s how the idea came from.

What is a Shuttle Vector

A shuttle vector is a type of plasmid vector constructed so that it can propagate in two different organisms such as prokaryotic and eukaryotic. (bacteria & yeast)

Shuttle vector is usually a plasmid vector.

Features of a Shuttle Vector

According to Kado and Tait, an ideal shuttle vector should have these features/characteristics:

• A shuttle vector must have the ability to replicate in many organisms
• Should be small enough to carry large foreign DNA
• Cloned genes should be easily detectable
• Should be nonpathogenic
• They must not induce any stress in the host

Let’s consider this as a shuttle vector. So, this must have origin of replication, selectable marker, multiple cloning site, right? As per definition, this must propagate in both bacteria and yeast. So, we need two origin of replication, one for bacteria and one for yeast. Then we also need two selectable markers, one for bacteria and one for yeast. Extra sequences for yeast host such as yeast centromeric sequence(CEN) or autonomous replicating sequence(ARS). Then we need multiple cloning site where our target gene will be inserted. This is common for both hosts.

Here, using bacterial ori and selectable marker it can propagate in bacterial host and using yeast ori and selectable marker it can propagate in yeast host. So, this is a structure of a shuttle vector.

Here are some examples of shuttle vector: Adenovirus shuttle vector, pMVD, pHV14, pEB10, pHP3, pJDB219, YEp etc.

Now we will see How to construct a shuttle vector.

Construction of a Shuttle Vector

This is a pKK223-3 plasmid that is 4.6kb in size. It has pBR322 ori, ampicillin resistance gene which is bla. And BamH1 Acc1 cutting site. pKK223-3 plasmid is digested with BamH1 and Acc1 to acquire a 2.7kb pKKD fragment consisting of pBR322 ori and bla (ampicillin resistance gene).

Now we need another ori for eukaryotes, right? pMVSCS1 is a eukaryotic plasmid that has pMVSCS1 ori and four antibiotic resistance gene. Now we need to add this ori into pKKD plasmid. To do that two plasmids (pKKD & pMVSCS1) are digested with XhoII and BamH1 and then ligated to construct an 8.3kb shuttle vector pMVD.

Here, pMVD has two ori and four antibiotic resistance genes against chloramphenicol (ClaI), sulfonamide (sulII), streptomycin (strAB) and ampicillin (bla).

Now what are the advantages and disadvantages of a shuttle vector.

Advantages:

• The biggest advantage of shuttle vectors is that they can propagate or can be manipulated in both host systems. They can be grown in E.coli and then can be used in a system which is more difficult or slower to use (yeast)
• Shuttle vectors can be used for both expression purpose and cloning purpose

Disadvantages:

• Carrying capacity is low (not larger than 20kb)

So, that’s it. I hope you understand the topic.

Thank you.

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