Manipulating DNA
Adapted from Biology by Ken Miller and Joe Levine, Prentice Hall, 2013
Until very recently, animal and plant breeders could not modify the genetic code of living things. They
were limited by the need to work with the variation that already exists in nature. Even when they tried to
add to that variation by introducing mutations, the changes they produced in the DNA were random and
unpredictable. Imagine, however; that one day biologists were able to go right to the genetic code, rewrite
an organism’s DNA, and make any changes they wanted. Imagine that biologists could swap genes at
will from one organism to another, designing new living things to meet specific needs. That day, as you
may know from scientific stories in the news, is already here.
How are changes made to DNA? Scientists use their knowledge of the structure of DNA and its chemical
properties to study and change DNA molecules. Different techniques are used to extract DNA from cells,
to cut DNA into smaller pieces, to identify the sequence of bases in a DNA molecule, and to make
unlimited copies of DNA. Understanding how these techniques work will help you develop an appreciation
for what is involved in genetic engineering.
THE TOOLS OF MOLECULAR BIOLOGY
Suppose you had a computer game you wanted to change. Knowing that the characteristics of that game
are determined by a coded computer program, how would you set about rewriting parts of the program?
To make such changes, a software engineer would need a way to get the program out of the computer,
read it, make changes in it, and then put the modified code back into the game.
Genetic engineering, making changes in the DNA code of a living organism, works almost the same way.
DNA extraction: How do biologists get DNA out of a cell? DNA can be extracted from most cells by a
simple chemical procedure: The cells are opened and the DNA is separated from the other cell parts.
Cutting DNA: DNA molecules from most organisms are much too large to be analyzed, so biologists cut
them precisely into smaller fragments using restriction enzymes. Hundreds of restriction enzymes are
known, and each one cuts DNA at a specific sequence of nucleotides. Restriction enzymes are
remarkably precise. Like a key that fits only one lock, a restriction enzyme will cut a DNA sequence only
it if matches the sequence precisely.
Separating DNA: How can DNA fragments be separated and analyzed? One way, a procedure known as
gel electrophoresis, can separate fragments of molecules according to their size. In gel electrophoresis,
a mixture of DNA fragments is placed at one end of a porous gel, and an electric voltage is applied to the
gel. When the power is turned on, DNA molecules, which are negatively charged, move toward the
positive end of the gel. The smaller the DNA fragment, the faster it moves. Gel electrophoresis can be
used to compare the genomes of different organisms or different individuals. It can also be used to locate
and identify one particular gene out of the millions of genes in an individual’s genome.
Restriction enzymes like the one shown here at left (EcoRI)
cut DNA strands at specific sites called recognition sites. EcoRI
recognizes the sequence GAATTC on any DNA strand and cuts
it between the G and the A, leaving “sticky ends” of DNA
fragments.
Gel Electrophoresis of DNA fragments
Gel electrophoresis is used to separate DNA
fragments. First, restriction enzymes cut
DNA into fragments. The DNA fragments
are then poured into wells on a gel, which
is similar to a thick piece of gelatin. A
electric voltage moves the DNA fragments
across the gel. Because longer fragments of
DNA move through the gel more slowly,
they do not migrate as far across the gel as
shorter fragments of DNA. Based on size,
the DNA fragments make a pattern of
bands on the gel. These bands can then be
compared with other samples of DNA.
DNA Fingerprinting
From: NOVA: Background to Forensic DNA Analysis
See NOVA Video “The Killer’s Trail”, then read the following background information.
DNA fingerprinting, also called forensic DNA analysis, is considered by many to be the police
investigator's secret weapon, a means of building cases or reanalyzing crimes using tiny bits of cryptic
evidence. Indeed, since 1986, when DNA evidence first entered the courtroom, the technique has aided
in the prosecution or defense of hundreds of cases, and in the exoneration of dozens of people wrongly
convicted.
As the acceptance of DNA evidence in the courtroom has grown, so has its importance, for the simple
reason that physical evidence linking suspects to crimes is often very sparse. Sometimes the weight of an
entire case -- even the life of the individual on trial -- rides on just a few drops of blood or strands of hair.
But as the reinvestigation of the 1954 murder of Marilyn Sheppard shows, the quality of evidence can
dramatically affect the amount of information that evidence provides.
In general, forensic DNA analysts compare the genetic makeup of tissue samples in search of similarities
and differences among them. They do this not by comparing all of the DNA contained in each cell, but
instead by marking a small number of segments and then checking for the presence or absence of those
segments in each sample.
One of the most common techniques, called variable number tandem repeat (VNTR) analysis, isolates
DNA segments that all have the same sequence of repeating letters (ATCATCATC, for example). It
organizes these repeating segments according to length, marks segments of a few different lengths, and
then compares samples based on the presence or absence of same-length segments. Two samples that
have as many as ten or twelve of these segments in common have very little chance -- one in several
million -- of being from two different people. Because VNTR analysis relies on samples that contain
relatively long strands of intact DNA, the technique usually cannot be used to analyze tissues as old as
the bloodstains found in the Sheppard house. For decades-old samples, forensic analysts use instead a
variety of polymerase chain reaction (PCR) techniques, including DQA1. The DQA1 analysis focuses on
one tiny segment of the genome. This segment, the DQA locus, holds eight alleles, each of which codes
for a different protein, and six of which can be marked and used for forensic analysis.
In a DQA1 analysis, lab technicians compare samples with regard to the presence or absence of these
six DQA alleles. However, because the alleles come in a small number of combinations (just forty-two),
it's possible that a person could match a sample from a crime scene without being the source of that
sample. For this reason, DQA1 is typically used only to rule out suspects, and not implicate them.
Pre-AP Biology: DNA Fingerprinting Homework
Name________________
Prd._____Date_________
Read the article above titled: Manipulating DNA.
1. Describe the three main steps involved for scientists to manipulate DNA:
A.
B.
C.
2. What are restriction enzymes and where do they come from? What do they do in biotechnology
research?
3. What role does gel electrophoresis play in the study of DNA?
Go to the following website. Click through the animation and answer the
questions that follow. NOVA: Create a DNA Fingerprint
http://www.pbs.org/wgbh/nova/education/body/create-dna-fingerprint.html
1. Describe the process of DNA fingerprinting.
2. In what ways is DNA fingerprinting like actual fingerprinting? In what ways is it different?
3. How conclusive is the evidence of DNA fingerprinting?
4. Where is there possibility of error?
Go to the following website. Watch the video “The Killer’s Trail”. Then, read the
article titled DNA Fingerprinting above and answer the following questions:
http://www.pbs.org/wgbh/nova/education/body/forensic-dna-analysis.html
1. How was DNA evidence used to prove that Dr. Sam Sheppard did not murder his wife? Why
wasn't this evidence used when the case first went to court?
2. Why do you think the DQA1 test was chosen for DNA analysis in this case, instead of
another, more powerful genetic test?
3. If the blood trail left at the murder scene wasn't Marilyn's or Sam's, whose blood might it have
been?
4. If you were a juror on this trial, would you be convinced by the DNA evidence? Why or why
not?