Sample information

It was just casually walking across first base on a baseball field. Once captured it hung from a thread in the middle of the vile.

Methods

Our first PCR test was set up on 10/16/2025 and was a duplex reaction that used both the Wolbachia 16S and Arthropod CO1 primers.  The annealing temperature was 49 degrees C.

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

Our first Gel image was taken on 10/23/2025:

-was run at 125 volts for ~36 minutes

-used this DNA Ladder: New England Biolabs 1 kb Plus DNA Ladder for Safe Stains (product #N0559S)

Our second PCR reaction was set up on 10/30/2025, used the same Taq polymerase as the first PCR reaction, and:

-was a single reaction that used the Wolbachia_F and Wolbachia_R primers

-used an annealing temperature of 55 degrees Celsius

Gel image was taken on 11/6/2025 and:

-was run at 125 volts for ~21 minutes

-used the DNA Ladder: New England Biolabs 1 kb Plus DNA Ladder for Safe Stains (product #N0559S)

Results

Due to a false positive DNA in the water during the first test further testing should be done with more care and accuracy.

After completion of the second PCR(single) Test, our confidence has risen to a high. This is due to all controls are properly displayed, the two positive controls are displaying the correct bands, the presence of Wolbachia remained negative throughout both tests.