A plasmid vector has two genes in it, a gene for streptomycin resistance (strR) and another for kanamycin resistance (kanR). There is a restriction site inside of the strR gene that you will use to move in your gene of interest. After cutting both your vector and insert you mix them together in a test tube. You then transform your mixture into E. coli and plate your cells on a master plate without any antibiotics. After colonies appear, you pick and re-plate onto a streptomycin containing plate and onto a kanamycin containing plate. What do you expect to grow on the master plate without any antibiotics? What do you expect to grow on the streptomycin plate? What do you expect to grow on the
Molecular Techniques
Molecular techniques are methods employed in molecular biology, genetics, biochemistry, and biophysics to manipulate and analyze nucleic acids (deoxyribonucleic acid (DNA) and ribonucleic acid (RNA)), protein, and lipids. Techniques in molecular biology are employed to investigate the molecular basis for biological activity. These techniques are used to analyze cellular properties, structures, and chemical reactions, with a focus on how certain molecules regulate cellular reactions and growth.
DNA Fingerprinting and Gel Electrophoresis
The genetic makeup of living organisms is shown by a technique known as DNA fingerprinting. The difference is the satellite region of DNA is shown by this process. Alex Jeffreys has invented the process of DNA fingerprinting in 1985. Any biological samples such as blood, hair, saliva, semen can be used for DNA fingerprinting. DNA fingerprinting is also known as DNA profiling or molecular fingerprinting.
Molecular Markers
A known DNA sequence or gene sequence is present on a chromosome, and it is associated with a specific trait or character. It is mainly used as a genetic marker of the molecular marker. The first genetic map was done in a fruit fly, using genes as the first marker. In two categories, molecular markers are classified, classical marker and a DNA marker. A molecular marker is also known as a genetic marker.
DNA Sequencing
The most important feature of DNA (deoxyribonucleic acid) molecules are nucleotide sequences and the identification of genes and their activities. This the reason why scientists have been working to determine the sequences of pieces of DNA covered under the genomic field. The primary objective of the Human Genome Project was to determine the nucleotide sequence of the entire human nuclear genome. DNA sequencing selectively eliminates the introns leading to only exome sequencing that allows proteins coding.
A plasmid vector has two genes in it, a gene for streptomycin resistance (strR) and another for kanamycin resistance (kanR). There is a restriction site inside of the strR gene that you will use to move in your gene of interest. After cutting both your vector and insert you mix them together in a test tube. You then transform your mixture into E. coli and plate your cells on a master plate without any antibiotics. After colonies appear, you pick and re-plate onto a streptomycin containing plate and onto a kanamycin containing plate.
- What do you expect to grow on the master plate without any antibiotics?
- What do you expect to grow on the streptomycin plate?
- What do you expect to grow on the kanamycin plate?
- How do you determine which colonies contain your gene of interest?
- As mentioned above, there is a restriction site found within the strR Please draw or write out two different examples of what restriction sites may look like. These should be short pieces of double stranded DNA.
- Using those same pieces of short DNA that you created above, show me what one of them may look like if it was cut with a blunt end restriction endonuclease. With the other example, show me what it would look like if it was cut with a sticky end restriction endonuclease.
- Why do restriction endonucleases exist in bacterial cells? What must they do to protect their own DNA?
- What are the pro’s and con’s of recognizing a short recognition sequence? What about a longer one?
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