UMAIR MASOOD Abbottābād Pakistan
What is a scientist?
- Someone who follows rules?
- Who publishes papers and gets grants?
- Can a scientist only do Art?
- Can a scientist never publish?
- Can a scientist never have worked at Harvard or MIT?
The term scientist conjures a very specific view in our minds. I don't want to be any of that. I can't be some of that.
The thing is to change what people think when they imagine a scientist they need someone to be that picture when they close their eyes.
That's you. The person who doesn't fit the stereotype of what a scientist is. You can be that scientist in someone's mind. People might have told you that you need school to be a scientist but the thousands of Biohackers across the world doing crazy shit would prove otherwise
Pocket PCR machine:
PCR is a method broadly used in molecular biology to manufacture several copies of a specific DNA segment
Pocket PCR machine components:
- Power cable
- Sample holder
- Pcr reaction display screen
- Sample holder
- Tightening screw
Pocket Pcr protocol:
- Prepare your Pcr reaction
- Connect your pcr to a power cable
- The main menu choose setup by rotating knob and then press it
- The pcr protocol can be setup through a graphical representation
- After scrolling through all the given parameter set cancel menu is show choose set to store the setting in the memory
- Back in the main menu choose run pcr to the pcr reaction
- The pcr run can be end by rotating the knob press to stop will appear in the display
Engineer any bacteria to fluoresce protocol:
- label one sterile microtube “A”. Mark another “B”
- utilize a micropipette, add 250 μl of CaCl2 solution (50 mM) to each tube
- utilize a sterile plastic loop to shift one or more than one 3 mm bacterial colonies from the streak plate to the “A” tube
- Close the tube lid and place the tube on ice
- utilize a new sterile loop to shift a mass of bacteria to the “B“ tube
- add 10ul of pGREEN directly into your “A”
- come back the “A and B” tube to the ice. Incubate both tubes on ice for 15 minutes
- add 250 μl of Luria broth to the “A” tube
- Add 250 μl of Luria broth to the “B” tube
- Label 1 LB plate “+ PLASMID” and 1 LB/AMP plate “+ PLASMID”
- Label 1 LB plate “– PLASMID” and 1 LB/AMP plate “‐ PLASMID”
- put-on 100μl of cell suspension from the “B” tube to the “LB ‐ PLASMID
- put-on 100 μl of cell suspension from the “A” tube to the “LB + PLASMID
- utilize a new sterile loop for each plate, spread the suspensions evenly around the surface muffle Parafilm around your four plates
- place the four plates upside down in a 37 C incubator. Incubate for 24 hours.
- Remove plates from incubator and observe the growth
- Observe plates with handheld UV light
DNA Synthesis from Oligos protocol:
- Design oligos for your desire gene
- Dilute the oligos up to 100uM
- Kinase(catalyzes the shifting of phosphate group) the oligos.
- For Thermostable Taq Ligase(10-25 cycles)
- For T4 ligase(5-10 cycles)
- 95°C denature, 15 sec
- 45-55°C anneal, 30 sec
- 20°C ligate
DNA cloning is a molecular biology technique that assemble many identical copies of a piece of DNA. In a representative cloning experiment, a target gene is load into a circular piece of DNA called a plasmid.
Steps of DNA cloning:
- Cutting and pasting DNA
- Bacterial transformation
- Protein manufacture
Isolation of genomic DNA from mouse tails protocol:
- Put a 0.5-1.5 cm mouse tail in a 1.5 ml Eppendorf tube and store frozen until digestion
- To each tube, add 500 ml of tail prep solution in the following sequence:
- 50 mM Tris-HCl (pH8.0)
- 100 mM EDTA
- 100 mM NaCl
- 1% SDS
- 0.5 mg Proteinase K/ml
- Digest at 55 °C overnight
- Cool sample on the ice
- Add 0.5 ml phenol, mix, and rotate12,000 rpm x 10m at room temperature
- shift the aqueous phase to a new Eppendorf tube
- Add 0.5 ml phenol:chloroform:isoamyl alcohol mix, and rotate 12,000 rpm x 10m Repeat Step with 0.5 ml chloroform
- shift the aqueous phase to a new Eppendorf tube, add equal volume 95% ethanol. lightly. Turnaround the tube several times to precipitate DNA.
- Centrifuge 10-15m at 12,000 rpm.
- Remove the supernatant
- wash a pellet with 70% ethanol
- Dry the pellet
- Resuspend pellet in 200-400 ml TE.
Check Gene expression at Home kit:
The central dogma of molecular biology consists of two steps: translation and transcription. Transcription is a process of synthesis an RNA copy of a gene sequence using DNA that encode a color marker you will able to check the production of RNA
P51™ Molecular Fluorescence Viewer:
- Sample holder
- Cover lid
- Uv-light source
Material and reagents:
1. Nuclease free water
2. DNA A
3. DNA B
5. Heat source 37°C 6.
- Label a tube 1 to 4
- Tube1 will be your negative control
- Tube 2 will be references
- Tube 3 will be your experiment reaction
- Tube 4 will be your experiment reaction
- Add a DNA to each pellet in the strip
- Tube 2 and 3 Add DNA(A) 5 μL
- Tube 4 Add DNA(B) 5 μL
- Do not add a liquid to tube 1
- Adding a regent to each tube
- Water tube 1, 7 μL
- Kanamycin tube 3, 2 μL
- Water tube 2 and 4, 2 μL
- Incubate the sample at 37°C
- After 15min observe the tubes under P51™ Molecular Fluorescence Viewer