Unit 3 Chapter 19: Bio Engineering

Question 1

What is genetic engineering

Answer 1

changing the genotype of an organism by transferring genes from one species to another

Question 2

What is selective breeding

Answer 2

• used to domesticate animals and plants
• all you need is to be able to select and breed the next generation
• you can wait for useful mutations to arise by chance or you can cross breed 2 closely related species

Question 3

What is gene cloning

Answer 3

moving a single gene from one organism to another

Question 4

What are the 2 reasons to use genetic engineering to move genes around

Answer 4

1. to make an organism express a new phenotype

2. to understand how the gene works

Question 5

What do beta islet cells do in diabetes

Answer 5

• in pancreas
• sense high levels of glucose and release insulin which tells body to absorbs glucose
• glucagon is released when there is too little glucose

Question 6

What causes diabetes

Answer 6

beta islet cells are destroyed by an autoimmune rxn so now the body can’t regulate glucose level

Question 7

Steps to integrating a new gene into an organism

Answer 7

make gene, insert into plasmid and then transform into bacteria

Question 8

Since bacteria can’t splice, genes without introns are needed how to you make these

Answer 8

make a DNA copy of the mRNA which has already been spliced

Question 9

How do you make a DNA copy of already spliced mRNA

Answer 9

use reverse transcriptase enzyme from retrovirus

Question 10

What is the DNA copy called

Answer 10

cDNA

Question 11

What is a vector

Answer 11

plasmid you are using to carry the gene you are cloning

Question 12

How do you get the vector to carry your gene

Answer 12

cut plasmid so its linear, and then join end of cDNA to end of plasmid to make a circle

Question 13

How do we cut DNA

Answer 13

restriction enzymes

Question 14

How do restriction enzymes work

Answer 14

identify a stretch of about 6 bps of a sequence and cut both ends at this point

Question 15

What is a restriction site

Answer 15

sequence of DNA recognized by restriction enzymes

Question 16

What do restriction enzymes leave

Answer 16

ragged/overhanging/sticky ends

Question 17

What is the advantage of sticky ends

Answer 17

the overhang will try to bond to a complementary overhang which make ligation rxn more efficient since sticky ends stay together

Question 18

How does cDNA stick to plasmid vector

Answer 18

DNA ligase (phosphodiester bonds)

Question 19

What are the basic steps to gene cloning

Answer 19

• cut up plasmid vectors with restriction enzymes
• mix with cDNAs and some ligase
• mix that with bacteria so bacteria takes it up through transformation
• then we have bacteria each with a plasmid containing different cDNA

Question 20

How do we know which bacteria have been transformed and which haven’t

Answer 20

put a gene on the bacteria that will show a specific phenotype on transformed bacteria and look for it (usually antibiotic resistance gene)

Question 21

What are transformation markers

Answer 21

the special phenotype you look for

Question 22

What is a cDNA library

Answer 22

allowing bacteria to grow and divide creating a little colony because bacteria represents a library of all of the mRNA (in cDNA form that can be made in a cell)

Question 23

What is a DNA probe

Answer 23

if you know the amino acid of the protein encoded by the gene you are cloning you can make one

• it’s a piece of DNA that helps you find complementary DNA
• look for radioactivity to find out which colony the probe is stuck to

Question 24

What is an expression vector

Answer 24

plasmid that will let the gene be expressed

Question 25

How do we make the gene be expressed

Answer 25

give it a promoter and terminator sequence so that the bacterial RNA polymerase will recognize it as a gene -put it in an expression vector (same process as putting into regular vector)

Question 26

How do we know if our plasmid has the right number of inserts

Answer 26

gel electrophoresis

Question 27

How does gel electrophoresis work

Answer 27

DNA is negatively charged and will move through an electric field toward the positively charged electrode (cathode). If you force the DNA to move through a thick tangle of agarose fibers (long strings of sugars) the bigger pieces will have a greater tendency to get stuck and will move more slowly. So if we purify DNA from several transformed colonies of bacteria, run each on a gel, stain the DNA with a fluorescent dye, we can find the DNA that is the right size

Question 28

What is the location of the restriction site into which you cloned your gene

Answer 28

is between the promoter, which consists of the lac promoter and the lac operator (but not the genes they control), and a terminator

Question 29

True or False: lactose can regulate the gene

Answer 29

true

Question 30

True or False: cystic fibrosis is a mendellian trait

Answer 30

true

Question 31

is cystic fibrosis the most common autosomal disease among caucasians

Answer 31

YES, its recessive though

Question 32

What is cystic fibrosis characterized by

Answer 32

respiratory problems, digestive issues, sterility inability to get rid of mucus, saltiness, mucus too dry

Question 33

How can we figure out what causes cystic fibrosis

Answer 33

1. find mutated gene and figure out what it encodes 2.sequence of protein can help us figure out what gene does (gene mapping) gets you to a group of genes not a single one

Question 34

What are restriction fragment length polymorphisms (RFLPs)

Answer 34

- variations in the sequence of a restriction site (like alleles kinda) - presence of them in DNA results in presence or absence of a restriction site - they segregate in a simple mendellian fashion - we can assay the presence or absence of several restriction sites - can be mapped like any allele - result in new band lengths in PCR

Question 35

How do you find RFLPs

Answer 35

LPs by looking for changes in the size of resulting DNA fragments after digesting genomic DNA with restriction enzymes

Question 36

How do you test for the presence of a specific RFLP

Answer 36

polymerase chain reaction

Question 37

What is the polymerase chain reaction

Answer 37

a method of producing large quantities of a short stretch of DNA from a known template DNA molecule - uses DNA polymerase - uses taq polymerase that loves hot temps

Question 38

What is bioinformatics

Answer 38

finding genes using a computer

Question 39

What are expressed sequence tags ESTs?

Answer 39

another way to find genes | -look for cDNAs and find genes that encode them

Question 40

So what is cystic fibrosis caused by

Answer 40

- a mutation in the gene encoding a chloride channel that regulates the flow of chloride across the plasma membrane of epithelial cells. Specifically the flow of chloride into the lung - When chloride isn’t regulated correctly, mucus becomes sticky and bacteria are able to grow

Question 41

Is the cystic fibrosis allele beneficial

Answer 41

ya, heterozygous individuals for the allele are protected from typhoid fever (bacteria are not internalized efficiently)