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Biomiga HP Plasmid Mini Kit - PD1711-02

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Description :   

Introduction

Key to the kit is our proprietary DNA binding systems that allow the high efficient binding of DNA to our ezBindTM matrix while proteins and other contaminates are removed under certain optimal conditions. Nucleic acids are easily eluted with sterile water or Elution Buffer. The purified DNA is ready for downstream applications such as cloning/subcloning, RFLP, sequencing, and transfection of HEK293 cells.

This kit is especially designed for purifying plasmid DNA from endA+ strains such as HB101, JM101, TG1 or their derived strains.

Important Notes

The yield of plasmid DNA depends on the origin of the replication and the size of the plasmid. The protocols are optimized for high copy number plasmid purification. For low copy number plasmids, both the culture volume and the buffer volume need to be scaled up 3 to 5 times. Please contact our customer service for further information and reference the table below for the commonly used plasmids,

Plasmid

Origin

High copy

Low copy

pACYC

P15A

10-12

pSC101

pSC101

5

pSuperCos

pMB1

10-20

pBR322

pMB1

15-20

pUC

Muted pMB1

500-700

pGEMR

Muted pMB1

300-400

pBluescriptR

ColE1

300-500

Storage and Stability

Buffer A1 should be stored at 4 ℃ once RNase A is added. All other materials can be stored at room temperature. The Guaranteed shelf life is 18 months from the date of purchase.

Kit Contents

Catalog Number

PD1711-00

PD1711-01

PD1711-02

Preps

4

50

250

ezBindTM Columns

4

50

250

Buffer A1

1.2 mL

15 mL

70 mL

Buffer B1

1.2 mL

15 mL

70 mL

Buffer N1*

1.5 mL

20 mL

90 mL

DNA Wash Buffer**

2 mL

15 mL

3 x 24 mL

Buffer KB

2.4 mL

30 mL

135 mL

EndoClean Buffer

(Optional)

100 uL

1 mL

(PD1214)

5 mL

(PD1214)

RNase A

1.2 mg

1.5 mg

7.0 mg

*Buffer N1 contains chaotropic salts, wear gloves and protective eyewear when handling.

**Add 8mLPD1711-0060 mL (PD1711-01) or 96 mL (PD1711-02) 96-100% ethanol to DNA Wash Buffer before use. The Final ethanol is 80% (v/v)

Before Starting

Prepare all components and get all necessary materials ready by examining this instruction booklet and become familiar with each steps.

Important

  • RNase A: Spin down RNase A vial briefly. Add the RNase A solution to Buffer A1 and mix well before use.
  • Add absolute ethanol as instructed to DNA Wash Buffer before use.
  • Buffer B1 precipitates below room temperature, it is critical to warm up the buffer at 37 ℃ to dissolve the precipitates before use.
  • Keep the cap tightly closed for Buffer B1 after use.
  • Ensure the availability of centrifuge capable of 13,000 rpm.
  • Carry out all centrifugations at room temperature.
  • Buffer N1 contains a chaotropic salt, wear gloves and protective eyewear when handling.

Materials supplied by users

  • 96- 100% ethanol.
  • 1.5 mL microcentrifuge tubes.
  • High speed microcentrifuge.
  • ddH2O.

Trouble Shooting Guide

Low Yield

Poor Cell lysis.

  • Resuspend pellet thoroughly by votexing and pipeting prior adding Buffer B1.
  • Make fresh Buffer B1 if the cap had not been closed tightly. (Buffer B1: 0.2N NaOH and 1%SDS).

Low Yield

Bacterial culture overgrown or not fresh.

Grow bacterial 12-16 hours. Spin down cultures and store the pellet at -20 ℃ if the culture is not purified the same day. Do not store culture at 4 ℃ over night.

Low Yield

Low copy-number plasmid.

Increase culture volume (up to 10mL for Miniprep). Scale up the volume of buffers accordingly.

No DNA

Plasmid lost in Host E.coli

Prepare fresh culture.

Genomic DNA contamination

Over-time incubation after adding Buffer B1.

Do not vortex or mix aggressively after adding buffer B1. Do not incubate more than 5 minutes after adding Buffer B1.

RNA contamination

RNase A not added to Buffer A1.

Add RNase A to Buffer A1.

Plasmid DNA floats out of wells while running in agarose gel, DNA doesn't freeze or smell of ethanol

Ethanol traces not completely removed from column.

Make sure that no ethanol residual remaining in the silicon membrane before elute the plasmid DNA. Re-centrifuge or vacuum again if necessary. 


  • Model: PD1711-02
  • Manufactured by: Biomiga

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