Biotech Exam 2 BSC4422 FIU Test
Questions and All Correct Answers
2026 Updated.
CHAPTER 3: Identify & Clone Gene of Interest - Answer DNA Libraries
Define DNA Libraries - Answer Collection of cloned DNA fragments,
from a particular organism,
contained in plasmid vectors, within a host bacteria.
Screen the library to? - Answer Pick out different genes of interest.
2 Types of DNA Libraries: - Answer Genomic DNA Libraries
&
Complementary DNA Libraries (cDNA Libraries)
Genomic DNA Libraries (3) - Answer 1. Chromosomal DNA,
from tissue of interest,
is -isolated- & -digested-,
with a restriction enzyme which produces many fragments [that include the entire genome]
2. Plasmid Vectors digested with same enzyme^
3. Ligase used to ligate chromosomal DNA fragments within plasmid vector
Recombinant Plasmid Vectors used to? - Answer Transform Bacteria
(so each bacteria will contain a single recombinant plasmid)
Disadvantages of Genomic Libraries (3)? - Answer 1. Clones introns!!
In addition to exons, introns cloned!
Majority of genomic DNA in eukaryotes is Introns (NON-CODING PIECES of DNA)
2. Hard & Time-consuming
Many organisms have large genomes, so seeking gene of interest is difficult
3. Not provide information on all levels of gene expression
Description of cDNA Libraries? - Answer mRNA from tissues of interest is Extracted ~~~>>
,cDNA Libraries
How to make double stranded DNA from mRNA? (3) - Answer 1. Short *Linker* double
stranded DNA sequences, which contain restriction enzymes recognition sites ADDED to the
ENDS of the cDNA.
2. Cut with restriction enzymes, cut vector, & ligate fragments to create:
3. Recombinant vectors
Disadvantage of cDNA libraries (1) - Answer Can be difficult to make IF a source tissue (with
an abundant amount of mRNA for the gene) is not available.
Advantage of cDNA libraries (3) - Answer 1. Does not clone introns!
2. Create & Screen isolated genes (that are only expressed under certain conditions in a tissue)
3. Collection of actively expressed genes from isolated mRNA.
Library Screening:
COLONY HYBRIDIZATION (9) - Answer 1. Bacteria with recombinant DNA grown on agar plate
2. Nylon or Nitrocellulose filter placed over plate
3. Treat filter with alkaline solution
-DOES 2 THINGS:
Lyses Cells & Denatures DNA
4. Bake filter or UV exposure
5. Incubate filter with a PROBE that is tagged with either:
-Radioactive nucleotide or Fluorescent dye
6. HYBRIDIZATION: Probe binds by H bonding to complementary sequence on the filter
7. Filter washed (to remove excess unbound probe)
8. Filter exposed to either:
X-ray film or digital camera
to detect fluorescent probe
9. Use film or picture to compare to the OG agar plate to identify which colonies contained the
recombinant plasmid with gene of interest.
Colony Hybridization:
Type of probe used depends on? - Answer What is already known about the gene of interest
Library screening rarely results in cloning of full-length gene.
, What do scientists look for? (3) - Answer 1. Get small pieces of the gene
2. Scientist look for overlapping sequences
3. They look for START (AUG) and STOP codons to know when full-length of gene is obtained
Polymerase Chain Reaction (PCR) - Answer Developed in 1983 by Katy Mullis (1944-2019).
Technique:
Making copies or amplifying a specific sequence of DNA in a short period of time.
Process of PCR (4) - Answer 1. Target DNA amplified is added to a tube
2. Mixed with nucleotides (dATP, dCTP, dGTP, dTTP), buffer with MgCl2, and DNA polymerase
3. Add Paired set of Forward and Reverse Primers (short single stranded DNA oligonucleotides
(18-22 nucleotides long)).
4. Reaction tube placed in THERMOCYCLER
PCR Cycle - Answer Thermocycler takes DNA through series of reactions
3 Stages:
1. Denaturation- 94-96 C
2. Annealing- (Hybridization) 52-58 C; primers H bond with complementary bases at the
opposite ends of target sequence
3. Extension- (Elongation) 70-75 C; DNA polymerase copies target DNA
End of 1 Cycle, DNA is? - Answer Doubled
Cycles are repeated? - Answer 20-30 times
Advantages of PCR (2) - Answer 1. Amplify millions of copies from small starting material in
short time
2. Calculate # of copies of target DNA starting with 1 molecule using Equation: 2^N
where N is number of cycles
How many copies of DNA will 22 cycles generate? - Answer 2^22 = 4, 194, 304
Applications of PCR: (6) - Answer 1. Making DNA probes
2. Studying gene expression
3. Detection of -viral & bacteria- infections
4. Diagnosis of genetic conditions
Questions and All Correct Answers
2026 Updated.
CHAPTER 3: Identify & Clone Gene of Interest - Answer DNA Libraries
Define DNA Libraries - Answer Collection of cloned DNA fragments,
from a particular organism,
contained in plasmid vectors, within a host bacteria.
Screen the library to? - Answer Pick out different genes of interest.
2 Types of DNA Libraries: - Answer Genomic DNA Libraries
&
Complementary DNA Libraries (cDNA Libraries)
Genomic DNA Libraries (3) - Answer 1. Chromosomal DNA,
from tissue of interest,
is -isolated- & -digested-,
with a restriction enzyme which produces many fragments [that include the entire genome]
2. Plasmid Vectors digested with same enzyme^
3. Ligase used to ligate chromosomal DNA fragments within plasmid vector
Recombinant Plasmid Vectors used to? - Answer Transform Bacteria
(so each bacteria will contain a single recombinant plasmid)
Disadvantages of Genomic Libraries (3)? - Answer 1. Clones introns!!
In addition to exons, introns cloned!
Majority of genomic DNA in eukaryotes is Introns (NON-CODING PIECES of DNA)
2. Hard & Time-consuming
Many organisms have large genomes, so seeking gene of interest is difficult
3. Not provide information on all levels of gene expression
Description of cDNA Libraries? - Answer mRNA from tissues of interest is Extracted ~~~>>
,cDNA Libraries
How to make double stranded DNA from mRNA? (3) - Answer 1. Short *Linker* double
stranded DNA sequences, which contain restriction enzymes recognition sites ADDED to the
ENDS of the cDNA.
2. Cut with restriction enzymes, cut vector, & ligate fragments to create:
3. Recombinant vectors
Disadvantage of cDNA libraries (1) - Answer Can be difficult to make IF a source tissue (with
an abundant amount of mRNA for the gene) is not available.
Advantage of cDNA libraries (3) - Answer 1. Does not clone introns!
2. Create & Screen isolated genes (that are only expressed under certain conditions in a tissue)
3. Collection of actively expressed genes from isolated mRNA.
Library Screening:
COLONY HYBRIDIZATION (9) - Answer 1. Bacteria with recombinant DNA grown on agar plate
2. Nylon or Nitrocellulose filter placed over plate
3. Treat filter with alkaline solution
-DOES 2 THINGS:
Lyses Cells & Denatures DNA
4. Bake filter or UV exposure
5. Incubate filter with a PROBE that is tagged with either:
-Radioactive nucleotide or Fluorescent dye
6. HYBRIDIZATION: Probe binds by H bonding to complementary sequence on the filter
7. Filter washed (to remove excess unbound probe)
8. Filter exposed to either:
X-ray film or digital camera
to detect fluorescent probe
9. Use film or picture to compare to the OG agar plate to identify which colonies contained the
recombinant plasmid with gene of interest.
Colony Hybridization:
Type of probe used depends on? - Answer What is already known about the gene of interest
Library screening rarely results in cloning of full-length gene.
, What do scientists look for? (3) - Answer 1. Get small pieces of the gene
2. Scientist look for overlapping sequences
3. They look for START (AUG) and STOP codons to know when full-length of gene is obtained
Polymerase Chain Reaction (PCR) - Answer Developed in 1983 by Katy Mullis (1944-2019).
Technique:
Making copies or amplifying a specific sequence of DNA in a short period of time.
Process of PCR (4) - Answer 1. Target DNA amplified is added to a tube
2. Mixed with nucleotides (dATP, dCTP, dGTP, dTTP), buffer with MgCl2, and DNA polymerase
3. Add Paired set of Forward and Reverse Primers (short single stranded DNA oligonucleotides
(18-22 nucleotides long)).
4. Reaction tube placed in THERMOCYCLER
PCR Cycle - Answer Thermocycler takes DNA through series of reactions
3 Stages:
1. Denaturation- 94-96 C
2. Annealing- (Hybridization) 52-58 C; primers H bond with complementary bases at the
opposite ends of target sequence
3. Extension- (Elongation) 70-75 C; DNA polymerase copies target DNA
End of 1 Cycle, DNA is? - Answer Doubled
Cycles are repeated? - Answer 20-30 times
Advantages of PCR (2) - Answer 1. Amplify millions of copies from small starting material in
short time
2. Calculate # of copies of target DNA starting with 1 molecule using Equation: 2^N
where N is number of cycles
How many copies of DNA will 22 cycles generate? - Answer 2^22 = 4, 194, 304
Applications of PCR: (6) - Answer 1. Making DNA probes
2. Studying gene expression
3. Detection of -viral & bacteria- infections
4. Diagnosis of genetic conditions
