Page 391 - Carrahers_Polymer_Chemistry,_Eighth_Edition

354 Carraher’s Polymer Chemistry

Along with the production of insulin, many other medical uses have been achieved for recombi-
nant DNA. This includes the production of erythropoetin, a hormone used to stimulate production
of red blood cells in anemic people; tissue plasminogen activator an enzyme that dissolves blood
clots in heart attack victims; and antihemophillic human factor VIII, used to prevent and control
bleeding for hemophilia people. These three important genetically engineered proteins were all
cloned in hamster cell cultures.
Gene engineering is the basis of gene therapy where genes are removed, replaced, or altered
producing new proteins for the treatment of such diseases as muscular dystrophy, some cancers,

adenosine deaminase deficiency, cystic fibrosis, and emphysema.
10.9 DNA PROFILING

DNA profiling is also referred to as DNA fingerprinting and DNA typing. It is used in paternity

identifi cation, classification of plants, criminal cases, identification of victims, heredity (of living,
recently deceased, and anciently deceased), and so forth. DNA profiling is a tool that allows a com-

parison of DNA samples.
While about 99.9% of our DNA is alike, the 0.1% is what makes us individuals, and it is this 0.1%
that allows for the identification of us as individuals. Of interest, it is not the within the gene por-

tions that makeup our different physical and mental characteristics, but the DNA profi ling employs
DNA taken from what is referred to as the “junk DNA.” Identification generally occurs because of

the formation of different lengths of this junk DNA after appropriate treatment. This junk DNA
contains the same sequence of base pairs, but in different repeat numbers. Thus, the sequence

ATTCGG may appear four times, five times, six times, and so on. There are typically some statisti-
cal number of repeats. Other sequences such as GGCATCC and AATGCAAT also appear in some
statistical number of repeats. While each of us have these different run sequences, individually we
have unique run lengths of these different run sequences. These run sequences are called variable

number of tandem repeats or VNTRs. The repeat runs used for identification are generally from

specific locations within a chromosome. Enzymes “cut” the associated DNA at specifi c locations
leading to decreases in DNA molecular weight. In fact, these DNA chain length decreases are
apparent as bandshifts in DNA gels. The combination of the differences in decreased DNA chain
lengths becomes unique as results are obtained from different enzymes are accumulated. These
changes in the movement of DNA segments are then compared with results from different individu-

als and identification as to whether the individuals are the same or different. The identity results are
often given as some percentage or ratio.

There are two basic types of DNA profiling: one that uses PCR enzymes and the second that
employs the restriction fragment length polymorphism (RFLP) enzymes. The PCR approach uti-
lizes a sort of molecular copying process where a specific region is selected for investigation. The

PCR approach requires only a few nanograms of DNA. The DNA polymerase makes copies of DNA
strands in a process that mimics the way DNA replicates naturally within the cell. Segments are
selected for special study and the results used to identify the DNA pattern.
With the exception of identical twins, each individual has a DNA profile that is unique. As

previously noted, in excess of 99.9% of the more than 3 billion nucleotides in human DNA are the
same. But, for every 1,000 nucleotides there is an average of one site of variation or polymorphism.
These DNA polymorphisms change the length of the DNA fragments produced by certain restric-
tion enzymes. The resulting fragments are called RFLP. Gel electrophoresis is typically employed
to separate the sizes and thus create a pattern of RFLLPs. The number and size of the fragments is
used to create the DNA profi le.
Several steps are involved in creating the genetic fingerprint. First, a sample of cells is obtained

from a person’s blood, bone, semen, hair roots, or saliva. The individual cells from the sample are
split open and DNA isolated. The DNA is treated with restriction enzymes that cleave the DNA
strands at specifi c locations, creating fragments of varying lengths and composition. The resulting

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