Host–Vector System - Class 12 Biotechnology - Chapter 2 - Notes, NCERT Solutions & Extra Questions
Renews every month. Cancel anytime
Your personal doubt-solving assistant
Chatterbot AI gives you 100% accurate answers to your questions in an instant.
Extra Questions - Host–Vector System | NCERT | Biotechnology | Class 12
💡 Have more questions?
Ask Chatterbot AINCERT Solutions - Host–Vector System | NCERT | Biotechnology | Class 12
Describe the importance of host vector system in rDNA technology.
In recombinant DNA (rDNA) technology, the host-vector system is crucial for successful gene cloning and expression. A host is a living cell that accepts foreign DNA, and a vector is a carrier that introduces this DNA into the host. The system's effectiveness hinges on the compatibility between the host and the vector.
Host organisms, like Escherichia coli or yeast, provide necessary cellular machinery for DNA replication and protein expression. They must be efficient in replicating the vector-DNA complex without degrading it. Accurate selection of the host is vital for high yield and stability of the desired product.
Vectors, such as plasmids or bacteriophages, are engineered to include elements like an origin of replication, selectable markers, and unique restriction sites. This is fundamental for efficient DNA insertion, replication, and retrieval.
Overall, the host-vector system significantly impacts the efficiency of gene cloning processes, affecting both yield and purity of the genetically engineered products. Proper integration and function of this system are essential for advancements in genetics, biotechnology, and medicine.
What are the major characteristics of a vector?
Vectors are pivotal tools in recombinoident DNA technology, and their effectiveness relies on several essential characteristics:
1. Size: Vectors should be small in size to facilitate easy manipulation and entry into host cells.
2. Origin of Replication (ori): A critical feature, the ori ensures that the vector can autonomously replicate inside the host organism, independent of the host's chromosome.
3. Unique Restriction Sites: Vectors must contain specific recognition sequences for restriction enzymes. These sites allow for the precise cutting and insertion of the gene of interest. The fewer the restriction sites beyond those necessary, the less the risk of unwanted cuts.
4. Selectable Markers: Vectors typically carry selectable marker genes that confer resistance to antibiotics like ampicillin or tetracycline. This attribute is essential for the selection of successfully transformed cells, as only those containing the vector will survive in an antibiotic-containing environment.
What is plasmid and what are its different types?
A plasmid is a small, circular, double-stranded DNA molecule that is distinct from a cell's chromosomal DNA and capable of replicating independently. Plasmids are commonly found in bacterial cells, and also occur in some eukaryotes. They play a crucial role in recombinant DNA technology by serving as vectors for gene cloning and genetic modification.
Plasmids can be categorized into different types based on their functions and replication capabilities:
1. Fertility (F) plasmids: These plasmids contain genes that facilitate the transfer of plasmids between bacterial cells through a process known as conjugation.
2. Resistance (R) plasmids: Provide bacteria with resistance against antibiotics or heavy metals. These plasmids are of particular importance in medicine because they can spread antibiotic resistance genes among bacterial populations.
3. Col-plasmids: Carry genes that produce bacteriocins, proteins that destroy or inhibit the growth of other bacteria, giving the host bacteria a competitive advantage.
4. Degradative plasmids: Encode enzymes that enable the digestion of unusual substances, such as toluene or salicylic acid, allowing bacteria to utilize these compounds as a source of carbon and energy.
5. Virulence plasmids: Carry genes that increase the virulence of the bacteria. These plasmids enable a bacterium to infect host tissues more effectively, producing toxins or other factors that enhance the bacterial infection process.
Discuss the strategy applied for the development of (pBR322) plasmid cloning vectors.
The development of the pBR322 plasmid vector signifies a vital advance in recombinant DNA technology. Originally derived from pBR313, a naturally occurring plasmid which was larger and less efficient, pBR322 was designed by making it more manageable and effective for laboratory use. Specifically, the strategy involved significantly reducing its size; from 9 kb in pBR313 to just 4,361 base pairs in pBR322. This reduction was achieved by eliminating the non-essential DNA sequences that did not contribute to its function as a vector. Importantly, pBR322 retained essential features like multiple unique restriction sites which allowed for versatile genetic manipulation and two antibiotic resistance genes, ampicillin (amp^R) and tetracycline (tet^R), which serve as selectable markers to identify successful recombinants. This smaller and more efficient vector became the most widely used vector for gene cloning due to its enhanced practical usability in the laboratory setting.
Briefly describe the structure of lambda bacteriophage and also discuss the role of lambda phage based vectors.
The lambda bacteriophage is a virus that infects E. coli bacteria and has a complex structure comprising a head (capsid) and a tail. The head contains the phage's linear double-stranded DNA genome, which becomes circular once inside the host due to cohesive ends known as "cos sequences".
Lambda phage based vectors, widely utilized in genetic engineering, exploit segments of the lambda phage genome. These vectors are designed for efficient DNA cloning, particularly in two broad classes: insertion vectors and replacement vectors. Insert not only integrates foreign DNA at a specific site within the phage genome but also ensures that non-essential genes can be replaced with desired DNA sequences, accommodating DNA segments up to 7.2 kb for insertion vectors and up to 20 kb for replacement vectors.
Lambda phage vectors have revolutionized molecular biology by enabling high-efficiency cloning and propagation of recombinant DNA, thereby providing potent tools for genetic research and biotechnology applications.
Discuss the M13 based vectors and its application.
M13-based vectors are derived from the filamentous bacteriophage M13, which infects Escherichia coli cells carrying the F plasmid. These vectors have a single-stranded DNA genome, typically around 6.4 kb in length. M13 vectors are particularly useful because they can produce single-stranded DNA, which is advantageous for applications such as sequencing and site-directed mutagenesis.
The life cycle of the M13 phage does not involve the lysis of the host cell; instead, the phage particles are secreted from the host without killing it. This property allows for continuous production of phage particles in cultures of E. coli. M13 vectors have been specifically engineered for cloning purposes. M13mp18, for example, is an M13 vector that facilitates blue/white selection of recombinant clones. This selection is critical in molecular cloning as it allows researchers to easily distinguish between recombinant and non-recombinant clones.
Applications of M13 vectors extend to phage display technology, where peptides or proteins are displayed on the surface of the phage and can be used for antibody production, vaccine development, or as a method for studying protein-protein interactions. Utilizing M13 vectors facilitates the rapid synthesis and testing of genetic modifications in a replicative form. These features make M13 vectors an essential tool in genetic research and biotechnology.
Differentiate between cosmids and phagemids.
Feature | Cosmids | Phagemids |
---|---|---|
Definition | Hybrid vectors resembling plasmids with lambda phage cos sites | Hybrid vectors combining traits of plasmids and phage |
Packaging | Can be packaged into phage coats using cos sites | Can form plaques or replicate as plasmids |
Origin | Derived from plasmids and specifically lambda phage cos sites | Derived from both plasmid and phage sequences |
DNA Insert Size | Typically up to 45 kbp | Smaller inserts compared to cosmids |
Uses | Useful for cloning large DNA fragments | Versatile, used in phage display or as cloning vectors |
Why is a vector required for cloning of a gene?
A vector is required for cloning a gene due to the following reasons:
Carrier and Propagation: A vector serves as a carrier for the desired DNA segment (insert) and facilitates its introduction into a host cell for propagation.
Autonomous Replication: Vectors have an origin of replication (ori), allowing them to replicate independently within a host organism.
Introduction of DNA into Host Cells: Vectors are typically designed to be easily incorporated into host cells.
Selection of Recombinants: Vectors usually contain selectable markers (e.g., antibiotic resistance genes) which enable researchers to identify and select cells that have successfully incorporated the vector and the gene of interest.
Therefore, without a vector, it would be challenging to manage, select, and propagate the desired DNA in a host organism for gene cloning applications.
A plasmid capable of getting integrated into host chromosome is called:
(a) Col plasmid
(b) Episome
(c) Ti plasmid
(d) R plasmid
The correct answer is:
(b) Episome
An episome is a type of plasmid that can integrate into the chromosomes of the host.
Why the replication of single copy plasmid called stringent replication?
The replication of single copy plasmids is termed stringent replication because their replication is tightly regulated and synchronized with the host cell's chromosomal DNA replication. This allows only one or a few copies of these plasmids per cell, ensuring that plasmid replication does not outpace the replication of the host's own genetic material. This control helps maintain plasmid stability and prevents it from overwhelming the host cell's resources, which could be detrimental to both the plasmid and the host.
Identify the incorrect match pair from the following:
(i) Multi copy plasmid | (a) Stringent replication |
(ii) Col plasmid | (b) Kills bacteria |
(iii) pBR322 | (c) Plasmid |
(iv) Prophage | (d) Phage genome inserted into a host genome |
The incorrect match pair from the table is:
(i) Multi copy plasmid - (a) Stringent replication
This is incorrect because high or multi copy plasmids undergo relaxed replication, not stringent replication, allowing for multiple copies of the plasmid per cell.
How can a large size eukaryotic gene insert be cloned?
For cloning a large size eukaryotic gene insert, special vectors like Yeast Artificial Chromosomes (YACs) are typically used. These are designed for cloning large DNA fragments (200-500 kb in size), which might include eukaryotic genes containing introns, potentially spanning hundreds of kilobases.
YACs consist of:
Two copies of a yeast telomeric sequence (telomeres are sequences at the ends of chromosomes)
A yeast centromeric sequence (helps in chromosome segregation during cell division)
A yeast autonomously replicating sequence (ARS) (where DNA replication begins)
Appropriate selectable markers
These components allow the YACs to function effectively in a eukaryotic system, ensuring that the large gene inserts are maintained and replicated within the host cells. This makes YACs ideal for managing and manipulating large DNA fragments characteristic of eukaryotic genes.
Assertion: An ideal vector should have selectable marker.
Reason: Selectable markers are required to screen out transformation.
(a) Both assertion and reason are true and the reason is the correct explanation of the assertion.
(b) Both assertion and reason are true but the reason is not the correct explanation of the assertion.
(c) Assertion is true but reason is false.
(d) Both assertion and reason are false.
(a) Both assertion and reason are true and the reason is the correct explanation of the assertion.
Explanation:
Assertion: An ideal vector should have a selectable marker. This is true as per the chapter content which explains that a vector needs to have a selectable marker to screen out transformants.
Reason: Selectable markers are required to screen out transformation. This is true and is the correct explanation for the assertion. They are essential for identifying cells that contain the vector, particularly those that have successfully taken up a recombinant vector incorporating the target gene.
Assertion: Cosmid is a hybrid vector.
Reason: Cosmid has properties of both plasmids and lambda phage vector.
(a) Both assertion and reason are true and the reason is the correct explanation of the assertion.
(b) Both assertion and reason are true but the reason is not the correct explanation of the assertion.
(c) Assertion is true but reason is false.
(d) Both assertion and reason are false
(a) Both assertion and reason are true and the reason is the correct explanation of the assertion.
Explanation:
Assertion: Cosmid is a hybrid vector. (True)
Reason: Cosmid has properties of both plasmids and lambda phage vector. (True)
Cosmids combine plasmid DNA features (such as replication function and selectable markers) with lambda phage properties (specifically the cos sites for DNA packaging). This makes the reason a correct explanation for why cosmids are considered hybrid vectors.
💡 Have more questions?
Ask Chatterbot AINotes - Host–Vector System | Class 12 NCERT | Biotechnology
Comprehensive Class 12 Notes on Host–Vector System in Recombinant DNA Technology
Introduction
Recombinant DNA technology has revolutionised the field of genetic engineering by allowing scientists to combine DNA from different sources. A successful application of this technology relies on a well-coordinated host–vector system. This article aims to provide comprehensive notes on the host–vector system, which is crucial for understanding the fundamentals of recombinant DNA technology.
Fundamentals of Host–Vector System
Key Components of Recombinant DNA Technology
Recombinant DNA (rDNA) technology involves two crucial steps:
- Isolation of the desired DNA (insert or target gene) from the source.
- Insertion of the target gene into a carrier DNA molecule known as a vector.
The vector containing the insert is known as recombinant DNA (rDNA). This rDNA is then introduced into an organism referred to as the host. The host–vector system is vital for gene cloning, expression, and manipulation.
Host in Gene Cloning
Requirements for a Suitable Host
A suitable host must allow the entry of rDNA, recognise it as self-DNA, and provide the necessary enzymes and proteins for smooth replication. Commonly used hosts include:
Prokaryotic Hosts
Escherichia coli (E. coli):
- Widely used due to its simple genetic structure and rapid growth.
- E. coli can double its population every 20 minutes, making it ideal for gene cloning. The K12 strain is particularly popular.
Bacillus subtilis:
- Used for its ability to secrete proteins encoded by cloned genes.
Eukaryotic Hosts
Yeast (Saccharomyces cerevisiae):
- Preferred for its eukaryotic cell structure and safety profile.
- Yeast can grow both sexually and asexually, facilitating various genetic manipulations.
Vectors in Gene Cloning
Essential Features of Vectors
For a molecule to act as a vector, it must meet several criteria:
- Small size for easy incorporation.
- An origin of replication (ori) for autonomous replication.
- Unique restriction sites to avoid fragmentation.
- Selectable markers for identifying transformants, e.g., antibiotic resistance.
Types of Vectors
Plasmids
Characteristics:
- Circular, double-stranded, extrachromosomal DNA.
- Capable of autonomous replication.
Development:
- High copy plasmids (replicate freely).
- Low copy plasmids (replicate along with chromosomal DNA).
Examples include plasmids like pBR322 and pUC19, commonly used in E. coli.
Bacteriophage Vectors
Lambda Phage:
- Infects E. coli and can insert large DNA fragments.
- Can enter lytic or lysogenic cycles for DNA replication.
M13 Bacteriophage:
- Consists of single-stranded circular DNA.
- Allows blue/white selection of recombinants.
Cosmids and Phasmids
Cosmids:
- Hybrid vectors combining plasmid and lambda phage elements.
- Capable of carrying large DNA inserts (up to 45 kbp).
Phasmids:
- True hybrid vectors combining plasmid and phage properties.
- Can replicate as either plasmid or phage.
graph TD;
A[Plasmid DNA] -->|Combines with| B[lambda Phage DNA];
B -->|Forms| C[Cosmid/Phasmid];
Advanced Vectors for Larger DNA Fragments
Yeast Artificial Chromosomes (YACs)
Function:
- Capable of carrying large DNA fragments (200-500 kbp).
Structure:
- Contains yeast telomeric sequences, a centromeric sequence, an autonomously replicating sequence (ARS), and selectable markers.
Bacterial and Phage Artificial Chromosomes (BACs and PACs)
Function:
- Suitable for cloning large fragments.
- BACs can insert DNA up to 300 kbp, while PACs handle sizes from 100-300 kbp.
Special Vectors
Expression Vectors
These vectors not only propagate the DNA insert but also ensure its expression. They contain:
- An efficient promoter for gene expression.
- Unique restriction sites for the insertion of the gene.
- A terminator sequence for the correct termination of transcription.
Shuttle Vectors
Designed to replicate in two different host types, shuttle vectors contain:
- Two origins of replication.
- Selectable markers for both types of hosts.
graph TD;
E[Prokaryotic Cell] -->|Hosts| F(Shuttle Vector);
F -->|Transfers to| G[Eukaryotic Cell];
Applications and Implications of Host–Vector Systems
Recombinant DNA technology using the host–vector system has numerous applications in:
- Biomedical Research: Creating genetically modified organisms for testing and therapy.
- Industrial Biotechnology: Producing enzymes, biofuels, and pharmaceuticals.
- Agricultural Biotechnology: Developing genetically modified crops with improved traits.
Conclusion
The host–vector system is a cornerstone of recombinant DNA technology. Understanding the characteristics and functions of various hosts and vectors is crucial for mastering the subject. This system opens the doorway to numerous applications, making it an indispensable tool in modern genetic engineering.
With these class 12 notes, you now have a thorough understanding of the host–vector system, setting a strong foundation for further studies and applications in recombinant DNA technology.
🚀 Learn more about Notes with Chatterbot AI