A new graduate student is performing a His-tag protein purification. They create a lysis buffer, wash...
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A new graduate student is performing a His-tag protein purification. They create a lysis buffer, wash buffer and elution buffer, with respective increasing concentrations of imidazole. You accidently use the elution buffer to lyse your cells, but decide to perform the His-purification anyways, and run your finished sample on an SDS-PAGE gel stained with instablue dye. What do you expect the end result to be and why
Homework Answers
Answer
If accidentally elution buffer has been used to lyse the cell then cell lysis will not happen no protein will come out of the cells so, at the end of the gell staining there will not be any protein band.
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A new graduate student is performing a His-tag protein
purification. They create a lysis buffer, wash...
A new graduate student is performing a His-tag protein purification. They create a lysis buffer, wash...
A new graduate student is performing a His-tag protein purification. They create a lysis buffer, wash buffer and elution buffer, with respective increasing concentrations of imidazole. You accidently use the elution buffer to lyse your cells, but decide to perform the His-purification anyways, and run your finished sample on an SDS-PAGE gel stained with instablue dye. What do you expect the end result to be and why (2)?
A graduate student has performed an in vitro pull-down assay to determine whether two proteins, Protein...
A graduate student has performed an in vitro pull-down assay to determine whether two proteins, Protein A and Protein B, interact with each other. Protein A is known to be 35 kDa and Protein B is 60 kDa. To determine whether there is an interaction the graduate student runs the sample down an SDS-PAGE gel, which is later stained with instablue dye. Unfortunately, the SDS, found in both the lysis buffer and SDS-PAGE gel, has expired (and is no longer...
A graduate student has performed an in vitro pull-down assay to determine whether two proteins, Protein...
A graduate student has performed an in vitro pull-down assay to determine whether two proteins, Protein A and Protein B, interact with each other. Protein A is known to be 35 kDa and Protein B is 60 kDa. To determine whether there is an interaction the graduate student runs the sample down an SDS-PAGE gel, which is later stained with instablue dye. Unfortunately, the SDS, found in both the lysis buffer and SDSPAGE gel, has expired (and is no longer...
a-d plz 7. Protein purification. As a graduate student, the first protein I purified was RNA...
a-d plz
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli- Some physical and chemical properties of Ecoli RNAP Molecular max470,000 g/mol polynentide compasitian (subunits): (50 kDa), k andok a pl = 5.34 substrates: NTPs cofactor: Mg? Purification protocol. E coli cells were broken usine Isozyme, yielding a cellualar extract containing a protcome solution 4M (NHOSO, was added to the cellular extract. A white protein precipitate was formed incuding RNAM...
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP)...
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli. Some physical and chemical properties of E. coli RNAP Molecular mass = 470,000 g/mol polypeptide composition (subunits): a (50 kDa), B (150 kDa) and 6 (70 kDa) pl = 5.34 substrates: NTPS cofactor: Mg Purification protocol. E. coli cells were broken using lysozyme, yielding a cellualar extract containing a proteome solution. 4M (NH4)2SO4 was added to the cellular extract. A white...
lab question 1. What is the basis of the different purification methods? 2. What are some...
lab question 1. What is the basis of the different purification methods? 2. What are some of the factors the might have interfered with your results? 3. How might you improve the process to increase the yield and purity? lab process E. coli BL21 (DE3) cells were transformed with the pET Topo-1521 vector containing a reading frame encoding the green fluorescent protein (GFP). Cells were cultured in M9ZB media at 37°C until the absorbance at 600 nm reached 0.7, at...
a-d plz
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli- Some physical and chemical properties of Ecoli RNAP Molecular max470,000 g/mol polynentide compasitian (subunits): (50 kDa), k andok a pl = 5.34 substrates: NTPs cofactor: Mg? Purification protocol. E coli cells were broken usine Isozyme, yielding a cellualar extract containing a protcome solution 4M (NHOSO, was added to the cellular extract. A white protein precipitate was formed incuding RNAM...
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli. Some physical and chemical properties of E. coli RNAP Molecular mass = 470,000 g/mol polypeptide composition (subunits): a (50 kDa), B (150 kDa) and 6 (70 kDa) pl = 5.34 substrates: NTPS cofactor: Mg Purification protocol. E. coli cells were broken using lysozyme, yielding a cellualar extract containing a proteome solution. 4M (NH4)2SO4 was added to the cellular extract. A white...
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