A plasmid map is the relative positions of the elements within the plasmid. A plasmid map can be read by understanding the features of the plasmid. They are the name and the size of the plasmid, the elements of the plasmid, their relative positions, and the orientation of the promoter. The origin of replication (ORI), antibiotic resistance gene, multiple cloning site (MCS), insert or gene of interest, promoter region, selectable marker, and primer binding site are the elements of a plasmid. The relative positions of the elements in a plasmid can be determined by mapping the plasmid. The process involved in the mapping of plasmids is called restriction mapping.
Key Areas Covered
1. What is a Plasmid Map
– Definition, Restriction Mapping
2. How to Read a Plasmid Map
– Features of a Plasmid MapP
Key Terms: Elements, Name and Size of the Plasmid, Orientation of the Promoter, Plasmid Map, Restriction Enzymes, Restriction Mapping
What is a Plasmid Map
A plasmid map is a graphical representation of a plasmid, which shows the locations of major landmarks or elements of the plasmid. The relative positions of elements within a plasmid can be identified by restriction mapping. A restriction map is a map of restriction recognition sites within a particular plasmid. Hence, it is involved in the digestion of the plasmid by restriction enzymes. A restriction map is shown in figure 1.
Figure 1: Restriction Map
Two methods can be used in restriction mapping:
1. Restriction Digestion
– Restriction digestion of the plasmid with one or two restriction enzymes
– Mapping the plasmid based on the sizes of the fragments resulted from the plasmid
2. Sequencing
– Sequencing the whole plasmid
– Identification of the elements of the plasmid
How to Read a Plasmid Map
Figure 2: Plasmid Map
The reading of a plasmid map mainly focuses on the features of the plasmid. They are;
1. The name and size of the plasmid
- The name and the size of the plasmid are indicated in the middle of the circular plasmid.
2. The elements of a plasmid –
- Origin of replication – the DNA sequence involved in the initiation of replication by recruiting the bacterial transcriptional machinery.
- Antibiotic resistance gene – allows the selection of plasmid-containing bacteria in a selective medium.
- Multiple cloning site – a short sequence that consists of several restriction recognition sites for the insertion of a foreign DNA fragment
- Insert – the gene of interest inserted into the plasmid
- Promoter region – binding site for RNA polymerase during transcription
- Selectable marker – allows the selection of successful expression of the inserted gene
- Primer binding site – serves as the initiation site for the PCR amplification of the plasmid for sequencing
3. The relative positions of the elements within the plasmid
- The relative positions of the plasmid elements are mapped by restriction mapping or sequencing.
4. The orientation of promoter
- The orientation of the promoter within the plasmid is important in determining the orientation of all the other elements of the plasmid, especially the orientation of the inserted gene. Transcription is initiated at the 3′ end of the promoter. Hence, the gene should be in the proper orientation in order to be expressed.
Conclusion
A plasmid map can be read by understanding the features of the plasmid map such as the name and the size of the plasmid, type of elements in the plasmid and their relative positions, and the orientation of the promoter.
Reference:
1. “What Is a Plasmid?” Addgene, Available here.
Image Courtesy:
1. “PDONR221 Map” By Nothingserious – Analyze Sequence: pDONR221. Addgene. Retrieved on 24 January 2016 (CC0) via Commons Wikimedia
2. “PGEX-3X cloning vector’ By Magnus Manske – Created by Magnus Manske (CC BY-SA 3.0) via Commons Wikimedia
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