Sample information

The arthropod was caught while Being dragged behind by another ant on a trail, the trail was dry and void of any moisture.

Methods

We used a DNA extraction protocol based on the insect adaptation of New England Biolabs’ Monarch Spin gDNA Extraction kit (Product # T3010).

The specimen was incubated for 30 minutes in a hot water bath at 56 degrees C, today’s PCR was a duplex reaction because we used both the Arthropod CO1 and Wolbachia 16S primers. I used an annealing temperature of 49 degrees Celsius. used Taq polymerase: New England Biolabs OneTaq Hot Start Quick-Load 2X Master Mix with Standard Buffer (#M0488S)

Gel electrophoresis system: Edvotek Gel Electrophoresis
Buffer: 1X TAE
DNA Stain: SYBR Safe

My first gel Image was taken on 10-21-25 and was run at 125 Volts for 30 minutes Using New England Biolabs 1 kb Plus DNA Ladder for Safe Stains (product # N0559S). This round told me that my arthropod tested false for Wolbachia

Our second PCR reaction was set up on 10-28-25, used the same Taq polymerase as the first PCR reaction, and:

was a Single.
We used both the Arthropod CO1 and Wolbachia 16S primer(s).

while using an annealing temperature of 55 degrees Celsius.

Second Round:

Gel electrophoresis system: Edvotek Gel Electrophoresis

Buffer: 1X TAE 

DNA Stain: SYBR Safe 

Protocol notes: ADD the following statements:

• Our second Gel image was taken on 11-4-25 and: 
  • was run at 125 volts for 30 minutes
  • used this DNA Ladder: New England Biolabs 1 kb Plus DNA Ladder for Safe Stains (product # N0559S)

Results: I am Highly positive that my arthropod is not Wolbachia infested, the reason for this is because after running the single Twice to check for any errors It came out with the same results witch leads me to Say in confidence that my arthropod test negatively.

This week we did DNA sequencing. For our arthropods only had the ZK-1 Come out due to the CD-1 arthropod being too distorted and messy.CO1: SequenceS16: Sequence

Results

After running the test the first instance showed that my arthropod was negative for Wolbachia, and after further testing the second round lead the same result. Giving me the ability to say in high confidence that my arthropod was indeed Wolbachia free. With also the Controls working. and we minimized any contamination from happening.