2. You want to clone gene A (1000 bp) into the SalI restriction site of the vector pBR322 illustrated below, pBR322 contains both ampicillin and tetracycline resistance genes. HindIII EcoRI EcoRV 4300 0 29 185 BamHI 375 4000 PstI 651 SalI 3607 tet amp 1000 pBR322 4361 bp 3000 ori 2296 2000 NdeI a) Which antibiotics would you include into the agar plates that would allow bacterial cells to grow that have been transformed with: i) No plasmid. ii) Non-recombinant pBR322. iii) Recombinant pBR322 with gene A cloned into the SalI site. b) Assume the cloning of gene A was successful, and you have isolated the recombinant plasmid. You set up separate restriction enzymes digestion for the non-recombinant and recombinant plasmids with (i) SalI and (ii) PstI + NdeI. Draw the gel profile you expect to observe when you subject the products of these four reactions to agarose gel electrophoresis. Indicate the sizes of all bands on the gel.
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You want to clone gene A (1000 bp) into the SalI restriction site of the vector pBR322 illustrated below, pBR322 contains both ampicillin and tetracycline resistance genes. a) Which antibiotics would you include into the agar plates that would allow bacterial cells to grow that have been transformed with: i) No plasmid. ii) Non-recombinant pBR322. iii) Recombinant pBR322 with gene A cloned into the SalI site. b) Assume the cloning of gene A was successful, and you have isolated the recombinant plasmid. You set up separate restriction enzymes digestion for the non-recombinant and recombinant plasmids with (i) SalI and (ii) PstI + NdeI. Draw the gel profile you expect to observe when you subject the products of these four reactions to agarose gel electrophoresis. Indicate the sizes of all bands on the gel.
Shaiju T.
Which antibiotics would you include into the agar plates that would allow bacterial cells to grow that have been transformed with: a) No plasmid. b) Non-recombinant pBR322. c) Recombinant pBR322 with gene A cloned into the SalI site. Assume the cloning of gene A was successful, and you have isolated the recombinant plasmid. You set up separate restriction enzyme digestions for the non-recombinant and recombinant plasmids with: a) SalI b) PstI c) NdeI Draw the gel profile you expect to observe when you subject the products of these four reactions to agarose gel electrophoresis. Indicate the sizes of all bands on the gel.
Sri K.
Describe the process of molecular cloning. a. The foreign DNA and plasmid are cut with the same restriction enzyme and DNA is inserted within the lacZ gene, whose product metabolizes lactose. The foreign DNA and vector are allowed to anneal. The vector is transferred to a bacterial host that is ampicillin sensitive and those with a disrupted lacZ gene show inability to metabolize X-gal. b. The foreign DNA and plasmid are denatured using high heat, and DNA is inserted within the lacZ gene, whose product metabolizes glucose. The foreign DNA and vector are allowed to anneal. The vector is transferred to a bacterial host that is ampicillin sensitive and disrupted lacZ gene will metabolize X-gal c. The foreign DNA and plasmid are cut with the same restriction enzyme and DNA is inserted randomly in the plasmid. The foreign DNA and vector are allowed to anneal. The vector is transferred to a bacterial host that is ampicillin sensitive and the disrupted lacZ gene shows inability to synthesize X-gal. d. The foreign DNA and plasmid are cut with the same restriction enzyme and DNA is inserted within the lacZ gene, whose product metabolizes lactose. The foreign DNA and vector are allowed to anneal. The vector is transformed into a viral host that is ampicillin sensitive and the disrupted lacZ gene show inability to synthesize X-gal.
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