This problem investigates issues encountered in sequencing the inserts in cDNA libraries.a. If you sequenced many clones individually, wouldn’tyou spend many of your resources inefficiently sequencing cDNAs for the same type of mRNA molecule over and over again? Explain. Does thisapparently inefficient process provide any useful information beyond the sequences of individual mRNAs?b. Suppose that you identified a clone with a cDNA insert that was 4 kb long. You could determine the entire sequence of the clone by shearing the DNA intosmall random fragments, cloning these fragments intoa vector to make a mini-shotgun library, and then sequencing hundreds of these clones to allow the computer to assemble the full sequence of the 4 kb–longinsert. However, this procedure would be inefficient.An alternative that requires many fewer sequencing reactions is called primer walking. This techniqueinvolves the synthesis of additional oligonucleotideprimers corresponding to cDNA sequences you havejust obtained. Diagram how you would sequence theentire 4 kb–long cloned cDNA using primer walking,indicating the vector and insert, all primers that youwould use, and all the sequences you would obtain.Assume that each sequence read is 1 kb.
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.
This problem investigates issues encountered in sequencing the inserts in cDNA libraries.
a. If you sequenced many clones individually, wouldn’t
you spend many of your
apparently inefficient process provide any useful information beyond the sequences of individual mRNAs?
b. Suppose that you identified a clone with a cDNA insert that was 4 kb long. You could determine the entire sequence of the clone by shearing the DNA into
small random fragments, cloning these fragments into
a vector to make a mini-shotgun library, and then sequencing hundreds of these clones to allow the computer to assemble the full sequence of the 4 kb–long
insert. However, this procedure would be inefficient.
An alternative that requires many fewer sequencing reactions is called primer walking. This technique
involves the synthesis of additional oligonucleotide
primers corresponding to cDNA sequences you have
just obtained. Diagram how you would sequence the
entire 4 kb–long cloned cDNA using primer walking,
indicating the vector and insert, all primers that you
would use, and all the sequences you would obtain.
Assume that each sequence read is 1 kb.
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