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Molecular biology services

eagle-i ID

http://eagle-i.itmat.upenn.edu/i/00000138-de4a-5e17-eb5b-b63c80000000

Resource Type

  1. Material production service

Properties

  1. Fee for service
    Yes
  2. Resource Description
    "<b>PCR, PCR clean up & Cloning</b> PCR of genomic DNA or BAC DNA is done using custom primers. Primers can also be designed at the facility. The optimization and the clean up of PCR products are done using various procedures. The cleaned up products can be directly processed for Sanger sequencing at the facility or used for cloning. <b>Transformation and Cloning</b> Design of cloning strategy followed by directional or non-directional clonings are performed using appropriate competent cells and vectors. Insertions sites are checked and orientations are determined by either restriction digestion and/or sequencing. PCR products are cloned using Invitrogen’s TA cloning system. <b>Sub-cloning</b> After designing the cloning strategy, DNA is sub-cloned into an appropriate expression vector followed by the preparation of the construct DNA for micro-injection into mouse zygotes. <b>Site-directed Mutagenesis</b> Mutagenesis including point mutation, insertion and deletion of single or multiple amino acids and large insertion are done using Stratagene’s Quick Change kit. This includes design and synthesis of the mutagenic primers, mutagenesis and verification by sequencing. The synthesis and the PAGE purification of the primers are outsourced to Invitrogen. <b>AMPure Bead Purification of Amplicons and DNA Fragments</b> Agencourt (Beckman) AMPure XP bead-based DNA purification of amplicons efficiently removes unincorporated dNTPs, primers, primer dimers, salts and other contaminants, resulting in high quality DNA suitable for sensitive downstream applications such as sequencing, genotyping, PCR, and sub-cloning. Recovery: 70 - 90% of double stranded and single stranded DNA with fragment sizes 150 bp - 10 kbp. This is the recommended purification method during sample preparation in most protocols for various new generation sequencing platforms."
  3. Service Fee URL
    http://www.med.upenn.edu/genetics/dnaseq/pricing.shtml
  4. Service Provided by
    Penn Genomics Analysis Core
  5. Website(s)
    http://www.med.upenn.edu/genetics/dnaseq/molbio.shtml
  6. Related Technique
    Site-directed mutagenesis
  7. Related Technique
    Polymerase chain reaction
  8. Related Technique
    Recombinant plasmid cloning
 
RDFRDF
 
Provenance Metadata About This Resource Record
  1. workflow state
    Published
  2. contributor
    ggrant (Gregory Grant)
  3. created
    2012-07-31T13:33:30.206-05:00
  4. creator
    fcoldren
  5. modified
    2020-03-09T12:15:31.255-04:00
Copyright © 2016 by the President and Fellows of Harvard College
The eagle-i Consortium is supported by NIH Grant #5U24RR029825-02 / Copyright 2016