Received blood, blood spot, and quill tip samples from Liam Berigan; University of Maine on 3-23-2022192-288 samples expected, most should be blood spots. This is a sponsored research project through Josh Jahner consisting of extractions and GBS preps.
Prepare Samples
Samples arrived 3-23-2022. Set1 and Set2
- Test extract some of the “Spare Feather” samples. In the large white box labeled “American WoodCock in the bottom of the hallway freezer, there is a small bag labeled “Spare Feathers” on top.
- Organize, label, and aliquot samples for extraction.
- Extract samples using Qiagen Blood and Tissue kit.
- Create sample submission form for extracted DNA samples.
- If there are a lot of samples above 150 ng/ul, normalize to 100 ng/ul on robot. If only a few, do by hand those few. If all, see if a universal dilution will get them all in range
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Extraction
All sample types will be extracted using Qiagen’s DNeasy Blood and Tissue Kits. The feather/quill samples will be extracted as shown in the Quill Tip Protocol involving an at least 12 hour digestion. SMin Quill and QLB samples showed high contamination after extraction. These samples were cleaned up using Zymo’s onestep-pcr-inhibitor-removal-kit. After quantification, a significant number of the QLB and quill samples were lower than the recommended range. We evaporated these and resuspended them in 50 ul ultra pure water.
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Create working pools. Load Pool A1 into columns 1-6 and A2 into columns 7-12 of the pooling plate.
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Pool
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Plate1 Col 1-6
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Plate1 Col 7-12
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Plate2 Col 1-6
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Plate2 Col 7-12
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1Smin Pool A1
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Pool
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Plate1 Col 1-6
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Plate1 Col 12-7
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Plate2 Col 1-6
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Plate2 Col 12-7
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1Smin Pool A2
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1
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2
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3
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4
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5
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6
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7
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8
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9
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10
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11
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12
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A
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PLT1_Col1
PLT1_Col7
PLT2_Col1
PLT2_Col7
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PLT1_Col2
PLT1_Col8
PLT2_Col2
PLT2_Col8
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PLT1_Col3
PLT1_Col9
PLT2_Col3
PLT2_Col9
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PLT1_Col4
PLT1_Col10
PLT2_Col4
PLT2_Col10
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PLT1_Col5
PLT1_Col11
PLT2_Col5
PLT2_Col11
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PLT1_Col6
PLT1_Col12
PLT2_Col6
PLT2_Col12
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PLT1_Col1
PLT1_Col12
PLT2_Col1
PLT2_Col12
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PLT1_Col2
PLT1_Col11
PLT2_Col2
PLT2_Col11
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PLT1_Col3
PLT1_Col10
PLT2_Col3
PLT2_Col10
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PLT1_Col4
PLT1_Col9
PLT2_Col4
PLT2_Col9
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PLT1_Col5
PLT1_Col8
PLT2_Col5
PLT2_Col8
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PLT1_Col6
PLT1_Col7
PLT2_Col6
PLT2_Col7
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B
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C
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D
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E
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F
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G
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H
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Pool
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Plate3 Rows A-B
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Plate3 Rows C-D
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Plate3 Rows E-F
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Plate3 Rows G-H
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PJDA
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Pool
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Plate3 Rows A-B
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Plate3 Rows D-C
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Plate3 Rows E-F
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Plate3 Rows H-G
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PJDB
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Pool
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Plate3
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Plate3
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Plate3
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Plate3
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Mix1
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A7, C7, E7, G7
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B7, D7, F7, H7
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Pool
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Plate3
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Plate3
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Plate3
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Plate3
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Mix2
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A7-D7
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E7-H7
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1
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2
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3
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4
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5
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6
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7
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8
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9
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10
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11
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12
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A
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PLT3_Row A, Row C, Row E, Row G
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PLT3_Row B, Row D, Row F, Row H
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PLT3_Row A, Row D, Row E, Row H
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PLT3_Row B, Row C, Row F, Row G
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PLT3_A7, C7, E7, G7
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B
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PLT3_B7, D7, F7, H7
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C
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PLT3_A7, B7, C7, D7
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D
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PLT3_E7, F7, G7, H7
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E
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F
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G
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PCR1:
Make MM1 in a 5mL tube:
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