Oly larvae RNA isolation
On 3/30/2018 I isolated total RNA from frozen larval samples. The larval samples were from my 2017 Oly OA study.
I used the RNAzol’s protocol for total RNA, RNAzol Total RNA Protocol; no issues encountered in the extraction process, however several resulted in very little RNA so I’m glad I did extra samples:
| Selected for Sequencing | RNA Sample # | Sample # | pH | Gonad Sex | Gonad Stage | Tissue Type | Tissue Sample date | RNA Extraction Date | RNA Concentration (ng/ul) | Approximate RNA isolated | Notes | |
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| X | 34 | 77-A | SN-6-Ambient | n/a | Frozen D-hinge larvae | 6/19/17 | SN-6 Ambient A | 3/30/18 | 61.2 | 1,163 | DNA also extracted | |
| X | 35 | 10-A | SN-6-Ambient | n/a | Frozen D-hinge larvae | 5/23/17 | SN-6 Ambient B | 3/30/18 | 71 | 1,349 | DNA also extracted | |
| 36 | 45-A | SN-6-Ambient | n/a | Frozen D-hinge larvae | 6/5/17 | SN-6 Ambient B | 3/30/18 | Out of range - TOO LOW | DNA also extracted | |||
| X | 37 | 69-A | SN-6-Ambient | n/a | Frozen D-hinge larvae | 6/15/17 | SN-6 Ambient B | 3/30/18 | 122 | 2,318 | DNA also extracted | |
| 38 | 37-A | SN-6-Ambient | n/a | Frozen D-hinge larvae | 6/3/17 | SN-6 Ambient B | 3/30/18 | 7.12 | 135 | |||
| X | 39 | 48-A | SN-6-Ambient | n/a | Frozen D-hinge larvae | 6/6/17 | SN-6 Ambient B | 3/30/18 | Out of range - TOO HIGH | |||
| 40 | 71-A | SN-6-Ambient | n/a | Frozen D-hinge larvae | 3/30/18 | Out of range - TOO LOW | ||||||
| X | 41 | 6-A | SN-10-Low | n/a | Frozen D-hinge larvae | 5/21/17 | SN-10-Low-B | 3/30/18 | 72.2 | 1,372 | DNA also extracted, but NOT from same sample vial (collected 2 vials from same larval release event) | |
| 42 | 62-A | SN-10-Low | n/a | Frozen D-hinge larvae | 3/30/18 | Out of range - TOO LOW | DNA also extracted | |||||
| 43 | 8-A | SN-10-Low | n/a | Frozen D-hinge larvae | 5/23/17 | SN-10-Low-B | 3/30/18 | Out of range - TOO HIGH | ||||
| X | 44 | 32-A | SN-10-Low | n/a | Frozen D-hinge larvae | 5/31/17 | SN-10 Low A | 3/30/18 | Out of range - TOO HIGH | |||
| X | 45 | 79-A | SN-10-Low | n/a | Frozen D-hinge larvae | 6/24/17 | SN-10 Low A | 3/30/18 | Out of range - TOO HIGH | |||
| 46 | 24-A | SN-10-Low | n/a | Frozen D-hinge larvae | 5/26/17 | SN-10 Low B | 3/30/18 | 88.8 | 1,687 | |||
| X | 47 | 26-A or 67-A | SN-10-Low | n/a | Frozen D-hinge larvae | 5/27/17 | SN-10 Low B | 3/30/18 | 194 | 3,686 | DNA also extracted - very little tissue left |
I also encountered black precipitate (see images below). Sam has encountered this before, and it didn’t seem to be a problem, check out his notebook post about it
I sent some of these (marked with an X under “selected for sequencing”) and some of the gonad samples extracted on March 27th off to Katherine for library prep and sequencing at the U-Chicago facility.
Written on March 30, 2018