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Lentiviral Mediated Gene Silencing in Human Pseudoislet Prepared in Low Attachment Plates
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JoVE 杂志 遗传学
Lentiviral Mediated Gene Silencing in Human Pseudoislet Prepared in Low Attachment Plates

Lentiviral Mediated Gene Silencing in Human Pseudoislet Prepared in Low Attachment Plates

DOI:
10.3791/59578-v

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11:24 min

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May 14, 2019

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, , , , ,
1Department of Internal Medicine, Carver College of Medicine,University of Iowa, 2Fraternal Order of Eagles Diabetes Research Center,University of Iowa, 3Roy J. Carver Department of Biomedical Engineering,University of Iowa

Chapters

  • 00:04Title
  • 00:42Overnight Culture of Human Islets for Recovery After Shipment
  • 01:32Preparation of Single Cell Suspension from Human Islets
  • 04:03Pseudoislet Formation and Transduction by Lentivirus
  • 08:44Results: Morphological and Functional Studies Using Gene Modified Human Pseudoislets
  • 09:53Conclusion

Summary

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A protocol to create gene modified human pseudoislets from dispersed human islet cells that are transduced by lentivirus carrying short hairpin RNA (shRNA) is presented. This protocol utilizes readily available enzyme and culture vessels, can be performed easily, and produces genetically modified human pseudoislets suitable for functional and morphological studies.

Tags

LentiviralGene SilencingHuman PseudoisletLow Attachment PlatesIslet FunctionGene Expression ModulationSHR8 TransductionProteolytic EnzymeCollagenolytic EnzymeCell DissociationCMRL MediumFetal Bovine Serum

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