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ALF_PLT1_2020_END

ALF_PLT2_2020_END

ALF_PLT3_2020_END

ALF_PLT4_2020_END

ALF_PLT5_2020_END

ALF_PLT6_2020_END

ALF_PLT7_2020_END

ALF_PLT8_2020_END

ALF_PLT1_2020_EPI

ALF_PLT2_2020_EPI

ALF_PLT3_2020_EPI

ALF_PLT4_2020_EPI

ALF_PLT5_2020_EPI

ALF_PLT6_2020_EPI

ALF_PLT7_2020_EPI

ALF_PLT8_2020_EPI

ALF_PLT9_2020_EPI

ALF_PLT1_2020_SOIL

ALF_PLT1_2017_GBS

ALF_PLT2_2017_GBS

ALF_PLT3_2017_GBS

ALF_PLT4_2017_GBS

ALF_PLT5_2017_GBS

ALF_PLT6_2017_GBS

ALF_PLT7_2017_GBS

ALF_PLT8_2017_GBS

ALF_PLT9_2017_GBS

ALF_PLT10_2017_GBS

ALF_PLT11_2017_GBS

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  • Equilibrate Beads to room Temperature

  • Add 24 ul of MagBeads to each well and 15 ul of replicate to same well of replicate

  • Pipette mix up and down 10 times.

  • Incubate at RT for 5 minutes

  • Secure plate on magnet plate; incubate at RT for 5 minutes (until wells are clear)

  • Remove 65 ul from each well; keep tips to left or right depending on the column to avoid bead pellet.

  • Add 100 ul Fresh 80% EtOH to each well. Incubate 30 seconds. Remove 100 ul from each well

  • Add 100 ul Fresh 80% EtOH to each well. Incubate 30 seconds. Remove 100 ul from each well

  • Reaspirate from each well to assure maximum EtOH removal

  • Allow plate to air dry for 7 minutes.

  • Remove sample plate from magnet plate.

  • Add 40 ul TE; pipette mix 10+ times. Incubate 2 minutes at RT.

  • Place sample plate back on magnet for 5 minutes or until all wells are cleared.

  • Transfer 40 ul to labeled transparent plate (Plate name_PCR_MIDs)

MasterMix (make for

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20 plates in

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50mL conical tube)

ul/rxn

Reagent

# of rxns

ul needed

3

5X Kapa HiFi Buffer

130021003900

6,300

0.45

10M dNTPs

13002100

585945

0.3

Kapa HiFi HotStart DNA Pol

13002100

390630

7.25

Ultra Pure H2O

130021009425

15,225

11

Total Volume

13002100

1423,300100

  • Add 11 ul to each well of a hard shell, full skirt plate. Seal with bubble strips and store in refrigerator until needed label “ALF PCR”. (If doing manually, one needs 2 ul of template and 2 ul of the primers).

Plates

16S MIDs

ITS MIDs

ALF_PLT9_2017_GBS

 long16S0A7

 longITS0A7

 long16S0B7

 longITS0B7

ALF_PLT10_2017_GBS

 long16S0C7

 longITS0C7

 long16S0D7

 longITS0D7

ALF_PLT11_2017_GBS

 long16S0E7

 longITS0E7

 long16S0F7

 longITS0F7

ALF_PLT9_2020_EPI

 long16S0G7

 longITS0G7

 long16S0H7

 longITS0H7

ALF_PLT1_2020_SOIL

 long16S0A8

 longITS0A8

 long16S0B8

 longITS0B8

Run on Thermocycler Program GSAF36:

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