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CopyControl™ BAC Cloning Kit (EcoR I)

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CopyControl™ BAC Cloning Kit (EcoR I)

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产品名称:

CopyControl™ BAC Cloning Kit (EcoR I)

货号
CCBAC1E
规格
1 Kit
存储温度
EpiLyse & EpiBlue Soln. are stored at room temp, the remainder is stored at -20℃
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产品描述

Applications

  • Preparation of BAC libraries that can be maintained at single copy number and induced to high-copy number as needed.

CopyControl™ BAC Cloning Kits* are complete kits containing all molecular biology reagents needed for constructing high-quality, large-insert Bacterial Artificial Chromosome (BAC) libraries. Each kit has enough reagents to make a complete human genome size library at 10X coverage. The kits feature the CopyControl™ pCC1BAC™ Vector which contains the E. coli F-factor single-copy origin of replication and oriV, a second origin of replication, which can be used to selectively induce to high copy number.

To maintain stability, CopyControl BAC clones are grown at very low copy number during the initial cloning and screening processes. Then, immediately before DNA purification, clones can be induced to 10-20 copies per cell within 2 hours by adding CopyControl Induction Solution to the culture, for higher yields of BAC DNA. The short, 2-hour induction of CopyControl BAC clones to high-copy number does not decrease their stability, based on analysis of Hind III restriction patterns of a large number of induced versus uninduced clones that ranged in size up to ~200 kb. Alternatively, the CopyControl BAC Autoinduction Solution can be used. Autoinduction Solution is added to the culture medium prior to inoculation, and also increases the cell growth leading to higher yield than standard Induction Solution. CopyControl BAC Cloning Kits reduce the time and labor required to construct a BAC library.

Phage T1-resistant TransforMax™ EPI300™-T1R Electrocompetent E. coli cells, or standard TransforMax™ EPI300™ Electrocompetent E. coli cells, which are required for induction of CopyControl BAC clones to 10-20 copies per cell, are available separately.

 

 

Benefits

  • Safely obtain higher DNA yields while maintaining the stability of single-copy-number clones.
  • Constructing BAC libraries is faster and easier.
  • CopyControl pCC1BAC cloning vectors are Cloning-Ready: linearized, dephosphorylated, and highly purified.
  • Cloning-Ready pCC1BAC vectors give >95% white colonies.
  • Estimate the size of the CopyControl BAC clones in as little as 4 hours without culture, Not I digests, or pulse-field gel electrophoresis.
  • CopyControl BACs are ideal for use in high-throughput (96-well) purifications when used with the CopyControl BAC Autoinduction solution.

Figure 3 (click to enlarge). The CopyControl™ BAC Cloning Kits significantly reduce the time and labor needed to construct a BAC library compared to standard methods.

Figure 1 Figure 1. DNA from induced (+) and uninduced (-) cultures of 4 CopyControl™ BAC clones. An equal number of cells were processed from both induced and uninduced cultures. Samples were resolved on a 0.8% agarose gel and stained with SYBR® Gold (Invitrogen). The induced cultures yielded >15-fold more CopyControl BAC clone DNA. M=BAC-Tracker™ Supercoiled DNA ladder. Figure 2 Figure 2. Not I digestion of 2 µl of DNA from induced (+) and uninduced (-) cultures of 4 CopyControl™ BAC clones. After resolving by PFGE and staining with SYBR® Gold (Invitrogen), clone sizes were: BAC1=130 kb, BAC2=125 kb, BAC3=140 kb and BAC4=145 kb. M=Pulsed-Field Gel Marker.

*Covered by issued and/or pending patents.
SYBR® is a registered trademark of Molecular Probes, Inc.

 

 

 

Catalog No. Size

CopyControl™ BAC Cloning Kit (BamH I)
   CCBAC1B 1 Kit
For constructing the equivalent of one 10X human BAC library.

CopyControl™ BAC Cloning Kit (EcoR I)
   CCBAC1E 1 Kit
For constructing the equivalent of one 10X human BAC library.

CopyControl™ BAC Cloning Kit (Hind III)
   CCBAC1H 1 Kit
For constructing the equivalent of one 10X human BAC library.

Contents: CopyControl™ pCC1BAC™ (BamH I, Hind III, or EcoR I) Cloning Ready Vector, Fast-Link™ DNA Ligase, Fast-Link™ 10X Ligation Buffer, ATP, BAC-Tracker™ Supercoiled DNA Ladder, EpiBlue™ Solution, EpiLyse™ Solution, Control Genomic DNA Insert, Control BAC Clone DNA (145 kb)

CopyControl™ pCC1BAC™ (BamH I Cloning-Ready) Vector
   CBAC311B 375 ng
Supplied at 25 ng/µl; vector is linearized at BamH I site and desphosphorylated.
EPI300™ cells must be purchased separately.

CopyControl™ pCC1BAC™ (EcoR I Cloning-Ready) Vector
   CBAC311E 375 ng
Supplied at 25 ng/µl; vector is linearized at EcoR I site and desphosphorylated.
EPI300™ cells must be purchased separately.

CopyControl™ pCC1BAC™ (Hind III Cloning-Ready) Vector
   CBAC311H 375 ng
Supplied at 25 ng/µl; vector is linearized at Hind III site and desphosphorylated.
EPI300™ cells must be purchased separately.

pCC1BAC/pIndigoBAC-5 Forward End-Sequencing Primer
   BFP0701 1 nmol

pCC1BAC/pIndigoBAC-5 Reverse End-Sequencing Primer
   BRP0801 1 nmol

CopyControl™ Induction Solution
   CCIS125 25 ml
1,000X concentrated solution. Filter sterilized.

CopyControl™ BAC Autoinduction Solution
   AIS107B 150 ml

 

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