Mosquitos

Sample information

Picture
Entry by: JC
Location
Collection date 10/06/2025
Captive / Cultivated? Wild-caught
Group Wolcott College Prep
Observations

long, thin, beige

Putative identification Arthropoda

Methods

Extraction kit DNeasy (Qiagen) blood and tissue kit
DNA extraction location Whole arthropod
Single or Duplex PCR Single Reaction
Gel electrophoresis system MiniPCR
Buffer TBE
DNA stain SeeGreen
Gel images
Protocol notes
  1. Place the gel tray containing your gel in the buffer chamber
    1. Ensure that the clear buffer chamber is inside the blueGel electrophoresis system.

 

  1. The wells of the gel should be on the same side as the negative electrode, away from the power button.

 

  1. Use a graduated cylinder to add 30 ml of 1X TBE electrophoresis buffer
    1. The buffer should just cover the gel and wells.

 

  1. Ensure that there are no air bubbles in the wells (shake the gel gently if bubbles need to be dislodged).

 

  1. Starting with the ladder in lane 1, gently hover your pipette tip over the well, and load 5 uL. Follow the loading key in the table below to load the rest of your gel. Remember to change tips between each sample. 
Lane Sample
1 DNA Ladder
2 Arthropod Sample #1
3 Arthropod Sample #2
4 Arthropod (+) Control (A #3)
5 Arthropod (-) Control (A #4)
6 (+) DNA Control (A #5)
7 (-) DNA Control (A #6)
8 Wolbachia Sample #1
9 Wolbachia Sample #2
10 Wolbachia (+) Control (W #3)
11 Wolbachia (-) Control (W #4)
12 (+) DNA Control (W #5)
13 (-) DNA Control (W #6)

 

  1. Place the orange cover on the blueGel electrophoresis system
    1. Match the positive and negative electrode signs on the orange lid with the corresponding positive and negative signs on the blue base.

 

  1. The orange lid should sit flush with the blue base using little force.

 

  1. Press the “Run” button
    1. Check that the green light beside the power button remains illuminated.

 

  1. Conduct electrophoresis for 20-25 minutes
    1. The colored dye should progress to about half the length of the gel.

 

  1. Longer electrophoresis times will result in better size resolution.

 

  1. Press the “light bulb” button to turn on the blueGel transilluminator.
    1. For best viewing, place the viewing chamber over the blueGel machine.

 

  1. Gels may be viewed at the end of the run or periodically throughout the run.

 

  1. Ensure that the bands in your gel have separated enough to clearly interpret your results
    1. Run the gel longer if needed to increase resolution.

 

  1. Take a picture and insert it in the correct post-lab question.

Results

Wolbachia presence No
Confidence level Medium
Explanation of confidence level

Some of the controls were not what they were supposed to be however some are correct.

Wolbachia 16S sequence
Arthropod COI sequence
Summary The Arthropoda was found to be negative for Wolbachia.
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