Extractingdna Essay, Research Paper

Extracting DNA from the Bacterium Escherichia coli

Introduction

Deoxyribonucleic acid is contained in all cells. The structure of DNA makes gene transmission

possible. Since genes are segments of DNA, DNA must be able to make exact copies of itself to enable the

next generation of cells to receive the same genes. The DNA molecule looks like a twisted ladder. Each

“side” is a chain of alternating phosphate and deoxyribose sugar molecules. The “steps” are formed by

bonded pairs of purine-pyrimidine bases. DNA contains four such bases the purines adenine (A) and guanine

(G) and the pyrimidines cytosine (C) and thymine (T).

The RNA molecule, markedly similar to DNA, usually consists of a single chain. The RNA chain

contains ribose sugars instead of deoxyribose. In RNA, the pyrimidine uracil (U) replaces the thymine of

DNA.

DNA and RNA are made up of basic units called nucleotides. In DNA, each of these is composed of a

phosphate, a deoxyribose sugar, and either A, T, G, or C. RNA nucleotides consist of a phosphate, a

ribose sugar, and either A, U, G, or C.

Nucleotide chains in DNA wind around one another to form a complete twist, or gyre, every ten

nucleotides along the molecule. The two chains are held fast by hydrogen bonds linking A to T and C to G

A always pairs with T (or with U in RNA); C always pairs with G. Sequences of the paired bases are the

foundation of the genetic code. Thus, a portion of a double-stranded DNA molecule might read: A-T C-G G-C

T-A G-C C-G A-T. When “unzipped,” the left strand would read: ACGTGCA; the right strand: TGCACGT.

DNA is the “master molecule” of the cell. It directs the synthesis of RNA. When RNA is being

transcribed, or copied, from an unzipped segment of DNA, RNA nucleotides temporarily pair their bases

with those of the DNA strand. In the preceding example, the left hand portion of DNA would transcribe a

strand of RNA with the base sequence: UGCACGU.

Genes and Protein Synthesis

A genetic code guides the assembly of proteins. The code ensures that each protein is built from

the proper sequence of amino acids.

Genes transmit their protein-building instructions by transcribing a special type of RNA called

messenger RNA (mRNA). This leaves the cell nucleus and moves to structures in the cytoplasm called

ribosomes, where protein synthesis takes place.

Cell biologists believe that DNA also builds a type of RNA called transfer RNA, which floats

freely through the cell cytoplasm. Each tRNA molecule links with a specific amino acid. When needed for

protein synthesis, the amino acids are borne by tRNA to a ribosome.

The Genetic Code

Experimental evidence indicates that the genetic code is a “triplet” code; that is, each series

of three nucleotides along the DNA molecule orders where a particular amino acid should be placed in a

growing protein molecule. Three-nucleotide units on an mRNA strand for example UUU, UUG, and GUU are

called codons. The codons, transcribed from DNA, are strung out in a sequence to form mRNA.

According to the triplet theory, tRNA contains anticodons, nucleotide triplets that pair their

bases with mRNA codons. Thus, AAA is the anticodon for UUU. When a codon specifies a particular amino

acid during protein synthesis, the tRNA molecule with the anticodon delivers the needed amino acid to the

bonding site on the ribosome.

The genetic code consists of 64 codons. However, since these codons order only some 20 amino

acids, most, if not all, of the amino acids can be ordered by more than one of them. For example, the

mRNA codons UGU and UGC both order cysteine. Because mRNA is a reverse copy of DNA the genetic code for

cysteine is ACA or ACG. Some codons may act only to signal a halt to protein synthesis. Since code

transmission from DNA to mRNA is extremely precise, any error in the code affects protein synthesis. If

the error is serious enough, it eventually affects some body trait or feature. In this study, DNA was

extracted from the bacterium Escherichia coli and then some of its physical properties were studied.

Methods & Materials

The DNA extraction was completed in one session of study. First, 5.0 ml. of E.coli suspension

medium was placed in a measuring cup and added to the tube of freeze-dried E.coli. The tube was then

capped very tightly and shook gently until the bacteria went into the suspension. Then, 1.0 ml. of sodium

dodecyl sulfate (SDS) was added to the E.coli suspension. The tube was then capped and was rotated gently

for over a period of five minutes. The suspension became more viscous as the bacteria was lysed. (Sodium

dodecyl sulfate (SDS), a detergent used in laundry products, removes the lipids from E.coli cell walls.

When the cell walls are damaged, the cells lyse, releasing the contents of into the E.cole suspension

medium.) The tube was placed for 30 minutes in a hot water bath preheated to 60-65 degrees C. The lysate

was removed from the water bath and was allowed to cool until it reached room temperature. With a pipet,

the cold ethanol was added to the spooling tube, while the spo!

oling rod was lowered into the E.coli suspension.

The spooling rod was slowly rotated in a continuous, clockwise direction. Fibers of the DNA came out of

the solution and attached to the glass rod, and the rod was turned a few more minutes until a visible

mass of DNA attached itself to it. The spooling rod was removed and immersed in 95% ethanol.

Results (see specimen)

Conclusion

Along with DNA, enzymes and many other proteins are present in the lysate. Enzymes harmful to DNA

are inactivated by heating the suspension to 60-65 degrees C,

a temperature that degrades proteins but not DNA. DNA must be heated to about 80 degrees C before it

denatures. DNA is also protected by sodium citrate, which was in the E.coli suspension medium. The

citrate ion is a resembling agent having a strong relationship for magnesium ions. These ions are

essential to the activity of DNAase, the enzyme that degrades DNA. DNA is soluble in water, but is

insoluble in ethanol. Thus, is the reason why the DNA fibers come out of the solution and attach to the

rod. DNA was then soaked in ethanol to stabilize it. Once the DNA was dried, it appeared white and

stringy, and there also was a considerable mass of DNA for so small of a sampling of bacteria. Thus

proving, that there is a great quantity of DNA in E.coli cells. It had great assymetry. Its length was

tremendous compared to its width. The length of the DNA molecule makes it very susceptible to splitting.

The DNA fibers can be fractured very easily, and because of its length, solutions !

of DNA are noticeable viscous.