Objective and summary:
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Run ethidium bromide gel electrophoresis for post-library prep samples F1 Juvenile Bay SCallop 8B 51 and 8B 52
- NOTE: I ran library prep and quantification tests for 8A1 and 8A2 yesterday and exhausted all extracted DNA from these samples. I also followed up with Qubit quantifcation postlibrary prep today along with Qubit for the raw sample 851, exhausted the remander of the extracted DNA for this sample
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Note: ran with a 1.5% agarose gel 100 mV for 60 minutes
Gel Electrophoresis
Gel map is as follows:
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‘raw’ refers to the samples run as direct aliquots of the DNA extraction using OMEGA EZNA extraction protocol here. Thus the sample ‘raw 8B 52’ is a positive control
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‘I’ refers to the Illumina DNA Prep protocol as suggested by manufacturers using all Illumina materials with 500 ng DNA samples SOP here
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‘8A 1 I’ refers to the Illumina DNA Prep protocol for library prep of sample 8A found here. We did not find a Qubit nor a gel reading, so the sample ‘8A 1 I’ is a negative control
| well | 1 | 2 | 3 | 4 | 5 |
|---|---|---|---|---|---|
| samp | L | raw 8B 52 | 8B 51 I | 8B 52 I | 8A 1 I |
Gel image
- we see strong high molecular weight band for the raw and a faint smear for bands in wells 3 and 4 (post library prep from today) These show a length of ~800-900 bp
Qubit quanitification
| Sample | DNA (ng/uL) | |
|---|---|---|
| RUN 1 | RUN 2 | |
| Standard | 150.85 | 160.50 |
| Standard | 14878.33 | 14492.81 |
| raw 8b 51 | 77.7 | 79.5 |
| 8B 51 I | 3.89 | 3.60 |
| 8B 52 I | 3.67 | 3.44 |
Summary
Takeaways from using Illumina DNA Prep library prep
- 500 ng input DNA yielded ~ 3.6 ng / ul output in 30 ul (~108 ng)
- size is roughly 800-900 bp in length
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adjustments to 6/6/2022 worked
- next step: (1) check whether this 5x decrease in DNA input is to be expected (2) also check whether this size range is to be expected