Difference between revisions of "Team:TU-Eindhoven/Project/Protocols"
| (18 intermediate revisions by 4 users not shown) | |||
| Line 8: | Line 8: | ||
<!-- | <!-- | ||
| − | |||
| − | |||
| − | |||
| − | |||
| − | |||
| − | |||
| − | |||
| − | |||
| − | |||
| − | |||
--> | --> | ||
</style></head> | </style></head> | ||
| Line 30: | Line 20: | ||
</body> | </body> | ||
<h1>General Protocols</h1> | <h1>General Protocols</h1> | ||
| − | <h6 | + | <h6><a href="https://static.igem.org/mediawiki/2017/e/ee/T--TU-Eindhoven--Protocols_General_Necessities.pdf" target="_blank">General Necessities</a> |
| − | Glycerol Stock<br/> | + | <br/><i>Preparation of antibiotics, growth media, stock solutions and more.</i><br/><br/> |
| − | Streaking Glycerol Stock<br/> | + | <a href="https://static.igem.org/mediawiki/2017/c/c2/T--TU-Eindhoven--Protocols_Glycerol_Stock.pdf">Glycerol Stock </a> |
| − | Nanodrop<br/> | + | <br/><i>Preparing the bacteria for long term storage in the -80 °C freezer.</i><br/><br/> |
| − | Agarose Gel Electrophoresis<br/> | + | <a href="https://static.igem.org/mediawiki/2017/6/6b/T--TU-Eindhoven--Protocols_Streaking_Glycerol_Stock.pdf" target="_blank">Streaking Glycerol Stock</a> |
| − | SDS-Page</h6><br/> | + | <br/><i>After storage in the -80 °C, the bacteria can be prepared for usage by streaking them on agar plates.</i><br/><br/> |
| + | <a href="https://static.igem.org/mediawiki/2017/d/d1/T--TU-Eindhoven--Protocols_Nanodrop.pdf" target="_blank">Nanodrop</a> | ||
| + | <br/><i>Nanodrop is used to determine the concentration of a DNA sample.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/7/7e/T--TU-Eindhoven--Protocols_Agarose_Gel_electrophoresis.pdf" target="_blank">Agarose Gel Electrophoresis</a> | ||
| + | <br/><i>Agarose Gel Electrophoresis is used to determine the length of DNA samples.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/7/7e/T--TU-Eindhoven--Protocols_Agarose_Gel_electrophoresis.pdf" target="_blank">SDS-Page</a> | ||
| + | <br/><i>SDS-Page is used to determine the size of a protein.</i></h6><br/> | ||
<h1>Traditional Cloning & BioBricking</h1> | <h1>Traditional Cloning & BioBricking</h1> | ||
| − | <h6>PCR Amplification<br/> | + | <h6><a href="https://static.igem.org/mediawiki/2017/d/d5/T--TU-Eindhoven--Protocols_PCR_Amplification.pdf" target="_blank">PCR Amplification</a> |
| − | PCR Purification<br/> | + | <br/><i>PCR Amplification is used to amplify a DNA sequence.</i><br/><br/> |
| − | Digestion<br/> | + | <a href="https://static.igem.org/mediawiki/2017/d/db/T--TU-Eindhoven--Protocols_PCR_Purification.pdf">PCR Purification</a> |
| − | Ligation<br/> | + | <br/><i>PCR Purification is used to purify the product from a PCR reaction.</i><br/><br/> |
| − | LIU PCR</h6><br/> | + | <a href="https://static.igem.org/mediawiki/2017/8/81/T--TU-Eindhoven--Protocols_Digestion.pdf">Digestion</a> |
| + | <br/></span><i>Digestion is used to cleave the plasmid and the insert in order to prepare them for ligation.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/6/69/T--TU-Eindhoven--Protocols_Ligation.pdf">Ligation</a> | ||
| + | <br/></span><i>Ligation is used to ligate a desired insert into a plasmid.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/b/bb/T--TU-Eindhoven--LIU_PCR.pdf">LIU PCR</a> | ||
| + | <br/></span><i>LIU PCR is used to insert additional amino acids in a DNA sequence.</i></h6><br/> | ||
<h1>Gibson Assembly</h1> | <h1>Gibson Assembly</h1> | ||
| − | <h6>Vector Linearization< | + | <h6><a href="https://static.igem.org/mediawiki/2017/e/ec/T--TU-Eindhoven--Protocols_Vector_Linearization.pdf">Vector Linearization</a> |
| − | < | + | <br/><i>Vector Linearization is used to linearize a vector, after which it can be used for Gibson Assembly.</i><br/><br/> |
| − | Gibson Assembly</h6><br/> | + | <a href="https://static.igem.org/mediawiki/2017/c/c0/T--TU-Eindhoven--Protocols_Gibson_Assembly.pdf">Gibson Assembly</a> |
| + | <br/><i>The NEBuilder HiFi Assembly protocol is used for Gibson Assembly to ligate gBlocks into a linearized vector.</i></h6><br/> | ||
<h1>Transformation, Protein Expression & Purification</h1> | <h1>Transformation, Protein Expression & Purification</h1> | ||
| − | <h6>Transformation into NovaBlue<br/> | + | <h6><a href="https://static.igem.org/mediawiki/2017/7/78/T--TU-Eindhoven--Protocols_Transformation_in_NovaBlue.pdf">Transformation into NovaBlue</a> |
| − | Plating<br/> | + | <br/><i>NovaBlue cells can be used for plasmid amplifcation.</i><br/><br/> |
| − | Colony Picking and Colony PCR<br/> | + | <a href="https://static.igem.org/mediawiki/2017/d/d2/T--TU-Eindhoven--Protocols_Plating.pdf">Plating</a> |
| − | Sequencing<br/> | + | <br/><i>Plating is required to amplify bacteria, by letting them grown on a growth medium.</i><br/><br/> |
| − | Miniprep<br/> | + | <a href="https://static.igem.org/mediawiki/2017/4/4c/T--TU-Eindhoven--Protocols_Picking_Colony_PCR.pdf">Colony Picking and Colony PCR</a> |
| − | Small culturing<br/> | + | <br/></span><i>Colony Picking and Colony PCR is used to evaluate if the inserted fragments have the correct length.</i><br/><br/> |
| − | Protein expression<br/> | + | <a href="https://static.igem.org/mediawiki/2017/7/7d/T--TU-Eindhoven--Protocols_Sequencing.pdf">Sequencing</a> |
| − | Protein Purification using a His-tag <br/> | + | <br/></span><i>Sequencing is used to determine the sequence of the amplified and transformed DNA.</i><br/><br/> |
| − | Protein Purification using a Strep-tag</h6><br/> | + | <a href="https://static.igem.org/mediawiki/2017/4/42/T--TU-Eindhoven--Protocols_miniprep.pdf">Miniprep</a> |
| + | <br/></span><i>Miniprep is used to obtain the plasmid DNA.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/0/0b/T--TU-Eindhoven--Protocols_Small_Culturing.pdf">Small culturing</a> | ||
| + | <br/></span><i>Small culturing is used to amplify a specific bacteria colony.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/3/37/T--TU-Eindhoven--Protocols_Protein_Expression.pdf">Protein expression</a> | ||
| + | <br/></span><i>This protocol is used to induce protein expression in bacteria from the small culture.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/e/e0/T--TU-Eindhoven--Protocols_Purification_His.pdf">Protein Purification using a His-tag</a> | ||
| + | <br/></span><i>This protocol is used to purify proteins using a His-tag.</i><br/><br/> | ||
| + | <a href="https://static.igem.org/mediawiki/2017/c/c3/T--TU-Eindhoven--Protocols_Purification_Strep.pdf">Protein Purification using a Strep-tag</a> | ||
| + | <br/><i>This protocol is used to purify proteins using a Strep-tag.</i> | ||
| + | </h6><br/> | ||
<h1>Testing</h1> | <h1>Testing</h1> | ||
| − | <h6> | + | <h6><a href="https://static.igem.org/mediawiki/2017/e/ec/T--TU-Eindhoven--Protocols_Flow_Chamber.pdf">Flow chamber</a> |
| − | Fluorescence | + | <br/><i>This protocol is used to prepare samples for microscopy.</i><br/><br/> |
| + | <a href="https://static.igem.org/mediawiki/2017/b/b6/T--TU-Eindhoven--Protocols_Measuring_Fluorescence.pdf">Measuring Fluorescence</a> | ||
| + | <br/><i>This protocol is used to measure the fluorescence of the fluorophores. It can also be used to measure FRET.</i><br/><br/> | ||
| + | <br/><i></i><br/> | ||
| + | </h6><br/> | ||
Latest revision as of 21:02, 1 November 2017
General Protocols
General Necessities
Preparation of antibiotics, growth media, stock solutions and more.
Glycerol Stock
Preparing the bacteria for long term storage in the -80 °C freezer.
Streaking Glycerol Stock
After storage in the -80 °C, the bacteria can be prepared for usage by streaking them on agar plates.
Nanodrop
Nanodrop is used to determine the concentration of a DNA sample.
Agarose Gel Electrophoresis
Agarose Gel Electrophoresis is used to determine the length of DNA samples.
SDS-Page
SDS-Page is used to determine the size of a protein.
Traditional Cloning & BioBricking
PCR Amplification
PCR Amplification is used to amplify a DNA sequence.
PCR Purification
PCR Purification is used to purify the product from a PCR reaction.
Digestion
Digestion is used to cleave the plasmid and the insert in order to prepare them for ligation.
Ligation
Ligation is used to ligate a desired insert into a plasmid.
LIU PCR
LIU PCR is used to insert additional amino acids in a DNA sequence.
Gibson Assembly
Vector Linearization
Vector Linearization is used to linearize a vector, after which it can be used for Gibson Assembly.
Gibson Assembly
The NEBuilder HiFi Assembly protocol is used for Gibson Assembly to ligate gBlocks into a linearized vector.
Transformation, Protein Expression & Purification
Transformation into NovaBlue
NovaBlue cells can be used for plasmid amplifcation.
Plating
Plating is required to amplify bacteria, by letting them grown on a growth medium.
Colony Picking and Colony PCR
Colony Picking and Colony PCR is used to evaluate if the inserted fragments have the correct length.
Sequencing
Sequencing is used to determine the sequence of the amplified and transformed DNA.
Miniprep
Miniprep is used to obtain the plasmid DNA.
Small culturing
Small culturing is used to amplify a specific bacteria colony.
Protein expression
This protocol is used to induce protein expression in bacteria from the small culture.
Protein Purification using a His-tag
This protocol is used to purify proteins using a His-tag.
Protein Purification using a Strep-tag
This protocol is used to purify proteins using a Strep-tag.
Testing
Flow chamber
This protocol is used to prepare samples for microscopy.
Measuring Fluorescence
This protocol is used to measure the fluorescence of the fluorophores. It can also be used to measure FRET.
