PCR adapter clean-up e5 samples

PCR adapter clean-up w/ e5 deep dive DNA samples

There are 3 deep dive samples (all POR) that keep failing the Zymo Pico prep (see previous notebook posts). There may be some adapter or inhibitor in the POR DNA that is preventing the preps from working. To combat this, we decided to use the Zymo PCR inhibitor remonval kit (see protocol here). We received 10 free samples of this kit.

I am going to clean the DNA extractions that I did (9/25/24) and that Kristina did (fall 2021) to see what will provide a better yield. Here is the sample info:

Number Species Timepoint Collection Date Site colony_id Extraction Date Extraction notebook post
421 Porites evermanni TP2 20200305 Site1 POR-82 20211008 https://kterpis.github.io/Putnam_Lab_Notebook/20211008-RNA-DNA-extractions-from-E5-project/
487 Porites evermanni TP2 20200305 Site1 POR-71 20211122 https://github.com/Kterpis/Putnam_Lab_Notebook/blob/master/_posts/2021-11-22-20211122-RNA-DNA-extractions-from-E5-project.md
489 Porites evermanni TP2 20200305 Site1 POR-79 20211129 https://github.com/Kterpis/Putnam_Lab_Notebook/blob/master/_posts/2021-11-29-20211129-RNA-DNA-extractions-from-E5-project.md
421 Porites evermanni TP2 20200305 Site1 POR-82 20240925 https://github.com/JillAshey/JillAshey_Putnam_Lab_Notebook/blob/master/_posts/2024-09-25-MiniprepPlus-DNA-extractions-E5.md
487 Porites evermanni TP2 20200305 Site1 POR-71 20240925 https://github.com/JillAshey/JillAshey_Putnam_Lab_Notebook/blob/master/_posts/2024-09-25-MiniprepPlus-DNA-extractions-E5.md
489 Porites evermanni TP2 20200305 Site1 POR-79 20240925 https://github.com/JillAshey/JillAshey_Putnam_Lab_Notebook/blob/master/_posts/2024-09-25-MiniprepPlus-DNA-extractions-E5.md

Protocol

  • Loosen filter cap and break bottom tip of filter off
  • Put filter in collection tube
  • Centrifuge for 3 mins at 8000g
  • Move filter into 1.5 mL tube and add sample
  • Centrifuge for 3 mins at 8000g

Qubit results

Used Qubit HS DNA kit

Number Extraction Date DNA1_ng/uL DNA2_ng/uL DNA average
421 20211008 0.586 0.574 0.58
487 20211122 0.967 0.941 0.954
489 20211129 0.81 0.798 0.804
421 20240925 0.117 0.114 0.1155
487 20240925 0.24 0.24 0.24
489 20240925 0.143 0.139 0.141
Written on October 14, 2024