Homework Answers
When the student will incorporate the DNA fragment using the Sall restriction enzyme (RE) then the DNA fragment will get inserted inbetween the tetracycline resistant(Tetr) gene because of the presence of Sall RE in between the Tetr gene. So this process will cause insertion inactivation where the Tetr gene will get disrupted and protein will not be made for Tetr and the cell will loose resistance to Tetr so when grown in media containing Tetracycline it will be neutralized.
Nutrient agar with ampicillin :- yes
Nutrient agar with tetracycline :- no
Nutrient agar with ampicillin and tetracycline :- no
NurtiNut agar with no antibiotics:- yes.
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A student clones a DNA fragment into the Sall site of pBR322. Sall is a restriction...
7. Explain the procedure for cloning DNA fragment into the plasmid PBR322 (shown on the right) (S...
7. Explain the procedure for cloning DNA fragment into the plasmid PBR322 (shown on the right) (S pts.). The gene fragment of interest was obtained by digestion of chromosomal DNA with the restriction enzyme Sall and subsequent purification using agarose gel electrophoresis. Which antiblotic would you use in the final step of the cloning procedure, and Pst why? EcoR Sal Ampicillin Tetracycline resistanica(Ter Amp) PBR322 4,361 bp) Origin of replicatiorn (ori Pvull 8. Assume that your gene fragment from question...
3. Below is a diagram of the pUC19 cloning vector (plasmid): Sall EcoRI Hinc Il Kpnl...
3. Below is a diagram of the pUC19 cloning vector (plasmid): Sall EcoRI Hinc Il Kpnl BamHI Accii Pst! Hind III T1 Sacl Xmal xbal BSDMI Sphi Smal Polylinker lacz -ori PUC19 amp Fig_18-08 Genetics, Second Edition 2005 WH Freeman and Company a. Describe briefly the purpose of the: ori: ampR; lacZt: Palxlinker b. When bacteria containing this plasmid are grown on media containing x-gal and ampicillin, why do sometimes you see blue colonies and sometimes white colonies? What does...
Restriction Mapping Below is a restriction map for the plasmid PGEN101 (total length - 20 kb)....
Restriction Mapping Below is a restriction map for the plasmid PGEN101 (total length - 20 kb). Using this map as a guide, give the number of restriction fragments along with their associated lengths that would result from digesting PGEN101 with the restriction enzymes EcoRI, BamHII, anda combination of EcoRI + BamHI. BamHI BamHI BamHI / PGEN101 (20 kb) Mb EcoRI Digest Performed Size Emments Obtained EcoRI........ BamHI.. EcoRI + BamHI.... Two freshmen college students, interested in becoming gene jocks, performed...
4. The DNA profiles to the right below represent four different individuals. o 111 Which of...
4. The DNA profiles to the right below represent four different individuals. o 111 Which of the following statements is consistent with the results? A. B is the child of A and C. B. C is the child of A and B. C. D is the child of B and C. D. A is the child of B and C. E. A is the child of C and D. <ULTE ILI UTILIT THITI TIL ILI 5. A eukaryotic gene with...
Cloning 2 Below is the restriction map of a 10 kb piece of DNA. Also shown...
Cloning 2
Below is the restriction map of a 10 kb piece of DNA. Also shown
below is a cloning vector which has two unique restriction enzyme
recognition sites, one for EcoRI (E) and one for HindIII (H). The
location of the kanamycin (kan) and ampicillin (amp) resistance
genes is also shown. Kanamycin and ampicillin are antibiotics that
are commonly used to select transformed E. colicells (consult the
Lab Manual for more information). Note that the HindIII site is
located...
The plasmid pFR55 is a useful vehicle for the cloning of any DBA sequence in E....
The plasmid pFR55 is a useful vehicle for the cloning
of any DBA sequence in E. coli. A restriction map of pFR55 showing
the restriction enzyme cutting sites is shown below. The plasmid
can replicate I'm E. coli and carries the tetracycline resistance
Tc and ampicillin resistance (Ap) genes for use as selectable
markers in E. coli.
a. you wish to insert a gene into the SalI site of
pFR55. How would you select for E. Coli cells that have...
Protein P is synthesized in relatively high amounts in the human pancreas. This protein has been...
Protein P is synthesized in relatively high amounts in the human pancreas. This protein has been isolated and purified, but its amino acid sequence has not been determined. We wish to clone the gene for protein P. (a) How can a probe be prepared to identify the gene for protein P? (b) If we have prepared a radioactive messenger RNA as our probe in part (a), how could we verify that it is the mRNA for protein P? (c) If...
Chromosomal and plasmid DNA can be cut into manageable pieces by restriction enzymes. Using agarose gel...
Chromosomal and plasmid DNA can be cut into manageable pieces by
restriction enzymes. Using agarose gel electrophoresis, the DNA
fragments can be separated on a gel, based on their lengths. In
order to see the fragments, a stain is typically added to the gel.
The size of each fragment can be determined by comparing each one
to a DNA molecular weight marker of known size.
Below is a map of pBR22 plasmid. The position and base pair
number of the...
If the target DNA (the 3.266 Kb E. coli genomic Bam HI fragment) has the same restriction sites on each end, there are t...
If the target DNA (the 3.266 Kb E. coli genomic Bam HI fragment)
has the same restriction sites on each end, there are two possible
orientations for the target DNA to insert into the plasmid. The
following restriction enzymes would cut the 3.266 kb Bam HI genomic
fragment containing the RecA gene once or twice or not at all. In
the tables below, list the expected DNA fragment sizes for the two
possible orientations. Round the DNA fragment sizes to...
Please answer these four questions. 6C. A type of plasmid used frequently as a vector in...
Please answer these four questions.
6C. A type of plasmid used frequently as a vector in recombinant DNA procedures carries gene markers for resistance to tetracycline (Tet') and to ampicillin (Amp). In one experiment, such plasmids were exposed to a restriction enzyme that cuts within the Amp' gene but leaves the Tet' gene intact. Plasmids exposed to the enzyme were next mixed with genomic restriction fragments from a cloned library of mouse DNA. The aim was to produce and detect...
7. Explain the procedure for cloning DNA fragment into the plasmid PBR322 (shown on the right) (S pts.). The gene fragment of interest was obtained by digestion of chromosomal DNA with the restriction enzyme Sall and subsequent purification using agarose gel electrophoresis. Which antiblotic would you use in the final step of the cloning procedure, and Pst why? EcoR Sal Ampicillin Tetracycline resistanica(Ter Amp) PBR322 4,361 bp) Origin of replicatiorn (ori Pvull 8. Assume that your gene fragment from question...
3. Below is a diagram of the pUC19 cloning vector (plasmid): Sall EcoRI Hinc Il Kpnl BamHI Accii Pst! Hind III T1 Sacl Xmal xbal BSDMI Sphi Smal Polylinker lacz -ori PUC19 amp Fig_18-08 Genetics, Second Edition 2005 WH Freeman and Company a. Describe briefly the purpose of the: ori: ampR; lacZt: Palxlinker b. When bacteria containing this plasmid are grown on media containing x-gal and ampicillin, why do sometimes you see blue colonies and sometimes white colonies? What does...
Restriction Mapping Below is a restriction map for the plasmid PGEN101 (total length - 20 kb). Using this map as a guide, give the number of restriction fragments along with their associated lengths that would result from digesting PGEN101 with the restriction enzymes EcoRI, BamHII, anda combination of EcoRI + BamHI. BamHI BamHI BamHI / PGEN101 (20 kb) Mb EcoRI Digest Performed Size Emments Obtained EcoRI........ BamHI.. EcoRI + BamHI.... Two freshmen college students, interested in becoming gene jocks, performed...
4. The DNA profiles to the right below represent four different individuals. o 111 Which of the following statements is consistent with the results? A. B is the child of A and C. B. C is the child of A and B. C. D is the child of B and C. D. A is the child of B and C. E. A is the child of C and D. <ULTE ILI UTILIT THITI TIL ILI 5. A eukaryotic gene with...
Cloning 2
Below is the restriction map of a 10 kb piece of DNA. Also shown
below is a cloning vector which has two unique restriction enzyme
recognition sites, one for EcoRI (E) and one for HindIII (H). The
location of the kanamycin (kan) and ampicillin (amp) resistance
genes is also shown. Kanamycin and ampicillin are antibiotics that
are commonly used to select transformed E. colicells (consult the
Lab Manual for more information). Note that the HindIII site is
located...
The plasmid pFR55 is a useful vehicle for the cloning
of any DBA sequence in E. coli. A restriction map of pFR55 showing
the restriction enzyme cutting sites is shown below. The plasmid
can replicate I'm E. coli and carries the tetracycline resistance
Tc and ampicillin resistance (Ap) genes for use as selectable
markers in E. coli.
a. you wish to insert a gene into the SalI site of
pFR55. How would you select for E. Coli cells that have...
Protein P is synthesized in relatively high amounts in the human pancreas. This protein has been isolated and purified, but its amino acid sequence has not been determined. We wish to clone the gene for protein P. (a) How can a probe be prepared to identify the gene for protein P? (b) If we have prepared a radioactive messenger RNA as our probe in part (a), how could we verify that it is the mRNA for protein P? (c) If...
Chromosomal and plasmid DNA can be cut into manageable pieces by
restriction enzymes. Using agarose gel electrophoresis, the DNA
fragments can be separated on a gel, based on their lengths. In
order to see the fragments, a stain is typically added to the gel.
The size of each fragment can be determined by comparing each one
to a DNA molecular weight marker of known size.
Below is a map of pBR22 plasmid. The position and base pair
number of the...
If the target DNA (the 3.266 Kb E. coli genomic Bam HI fragment)
has the same restriction sites on each end, there are two possible
orientations for the target DNA to insert into the plasmid. The
following restriction enzymes would cut the 3.266 kb Bam HI genomic
fragment containing the RecA gene once or twice or not at all. In
the tables below, list the expected DNA fragment sizes for the two
possible orientations. Round the DNA fragment sizes to...
Please answer these four questions.
6C. A type of plasmid used frequently as a vector in recombinant DNA procedures carries gene markers for resistance to tetracycline (Tet') and to ampicillin (Amp). In one experiment, such plasmids were exposed to a restriction enzyme that cuts within the Amp' gene but leaves the Tet' gene intact. Plasmids exposed to the enzyme were next mixed with genomic restriction fragments from a cloned library of mouse DNA. The aim was to produce and detect...
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