Background
Intake of Field Sourced Tunciates
Field sourced Botryllus schlosseri are brough from the City of De Moines Marina on a weekly basis. Botryllus schlosseri colonies are brought back from the field and kept in UW Tacoma’s recirculating seawater system (27 PPT, 17 C, 8 pH) until they meet the prerequistes to be entered in the nickel exposure study. Only individuals that are capable of withstanding the transition into the lab setting, have adhered a sufficient number of star-systems to the glass slide, and have entered stage C2 are utilized in the following exposures.
Field Collection Dates and Info
6 collections have occurred thus far for the purposes of using whole systems in nickel chloride exposures.
| Collection Date | Collection | Number of Genets Used in Study | Number Collected |
|---|---|---|---|
| 08-30-2024 | 12 | 3 | NA |
| 09-10-2024 | 13 | 2 | NA |
| 09-18-2024 | 14 | 6 | NA |
| 09-26-2024 | 15 | 3 | NA |
| 10-02-2024 | 16 | 0 | NA |
| 10-15-2024 | 17 | TBD | 9 |
Number of Days in Recirculating System Prior to Preservation
As Botryllus schlosseri sourced from the field cannot survive in the recirculating seawater system indefinitely, it is critical that the material is used prior to degradation. Degredation of the specimen typically occurs by the 3rd to 4th week in the lab. However, as we require the colony systems to be fully adhered to the slide prior to exposure start and for them to be the right blastogenic stage, we must wait for those factors to occur. The number of days that animals have spent in the system should be roughly the same for the purposes of this -omics study.
Below’s table separates the average days the individuals spent in lab and in the study from the day of collection to the day of tissue preservation. Roughly, animals spend 10 days in the recirculating system and in the exposure prior to freezing of the tissue.
| Average Days | SD | |
|---|---|---|
| Control | 9 | 3 |
| 100 mg/L Nickel Treatment | 12 | 2 |
| All Samples | 10 | 3 |
Sample Processing
The goal by the end of this study is to have 10 genets saved from the control and 10 genets saved from the 100 mg/L nickel exposure treatment. This would result in a total of 40 samples as tissue from each genet was divided for the fate of transcriptomics and proteomics.
Samples for the transcriptomics will be extracted for their RNA in-house and will be sent off to collaborators to evaluate quality on their bioanalyzer by mid-November.
Bioanalyzer Results for transcriptomics
| Sample ID | Bioanalyzer Name | Treatment | [Sample] ng/ul | Elution Volume (uL) | Date RNA Extracted | Bionalyzer RIN |
|---|---|---|---|---|---|---|
| 100 R1 9/11/24 | 100 R1 | 100 mg/L NiCl(II) | 0.8 | 40 | 11/29/2024 | failed |
| 100 R2 9/4/24 | 100 R2 | 100 mg/L NiCl(II) | 129.5 | 40 | 11/29/2024 | 7.3 |
| 100 R3 9/19/24 | 100 R3 | 100 mg/L NiCl(II) | 284 | 40 | 11/29/2024 | 7.2 |
| 100 R4 9/29/24 | 100 R4 | 100 mg/L NiCl(II) | 44.5 | 40 | 11/27/2024 | 7.3 |
| 100 R5 9/29/24 | 100 R5 | 100 mg/L NiCl(II) | 9.9 | 40 | 11/27/2024 | 10 |
| 100 R6 11/2/24 | 100 R6 | 100 mg/L NiCl(II) | 64.9 | 40 | 11/29/2024 | 9.6 |
| 100 R7 11/2/24 | 100 R7 | 100 mg/L NiCl(II) | 2.3 | 40 | 11/27/2024 | failed |
| 100 R8 11/15/24 | 100 R8 | 100 mg/L NiCl(II) | 78.6 | 40 | 11/29/2024 | 9.8 |
| 100 R9 11/17/24 | 100 R9 | 100 mg/L NiCl(II) | 1.3 | 40 | 11/27/2024 | failed |
| Ctrl R1 9/12/24 | C R1 | 0 mg/L NiCl(II) | 52.6 | 40 | 11/27/2024 | 7.4 |
| Ctrl R2 9/19/24 | C R2 | 0 mg/L NiCl(II) | 114.1 | 40 | 11/29/2024 | 7.3 |
| Ctrl R3 9/24/24 | C R3 | 0 mg/L NiCl(II) | 340.3 | 40 | 11/27/2024 | 9.7 |
| Ctrl R4 9/24/24 | C R4 | 0 mg/L NiCl(II) | 67.8 | 40 | 11/29/2024 | 7.4 |
| Ctrl R5 9/25/24 | C R5 | 0 mg/L NiCl(II) | 48.2 | 40 | 11/27/2024 | 10 |
| Ctrl R6 0/29/24 | C R6 | 0 mg/L NiCl(II) | 60.9 | 40 | 11/29/2024 | 9.3 |
| Ctrl R7 10/1/24 | C R7 | 0 mg/L NiCl(II) | 2.6 | 40 | 11/27/2024 | failed |
| Ctrl R8 10/9/24 | C R8 | 0 mg/L NiCl(II) | 38.1 | 40 | 11/29/2024 | 7.3 |
| Ctrl R9 10/9/24 | C R9 | 0 mg/L NiCl(II) | 116.3 | 40 | 11/29/2024 | 7.3 |
| Ctrl R10 10/15/24 | C R10 | 0 mg/L NiCl(II) | 22.1 | 40 | 11/27/2024 | 10 |
Seems like we will have enough of each treatment (best 6 from control anbd best 6 from treatment) to send off for RNAseq.