• Register
Forgot Password

If you have forgotten your password you can enter your email here and get a temporary password sent to your email.


Leaving Community

Are you sure you want to leave this community? Leaving the community will revoke any permissions you have been granted in this community.


Lentiviral vectors and protocols for creation of stable hESC lines for fluorescent tracking and drug resistance selection of cardiomyocytes.

BACKGROUND: Developmental, physiological and tissue engineering studies critical to the development of successful myocardial regeneration therapies require new ways to effectively visualize and isolate large numbers of fluorescently labeled, functional cardiomyocytes. METHODOLOGY/PRINCIPAL FINDINGS: Here we describe methods for the clonal expansion of engineered hESCs and make available a suite of lentiviral vectors for that combine Blasticidin, Neomycin and Puromycin resistance based drug selection of pure populations of stem cells and cardiomyocytes with ubiquitous or lineage-specific promoters that direct expression of fluorescent proteins to visualize and track cardiomyocytes and their progenitors. The phospho-glycerate kinase (PGK) promoter was used to ubiquitously direct expression of histone-2B fused eGFP and mCherry proteins to the nucleus to monitor DNA content and enable tracking of cell migration and lineage. Vectors with T/Brachyury and alpha-myosin heavy chain (alphaMHC) promoters targeted fluorescent or drug-resistance proteins to early mesoderm and cardiomyocytes. The drug selection protocol yielded 96% pure cardiomyocytes that could be cultured for over 4 months. Puromycin-selected cardiomyocytes exhibited a gene expression profile similar to that of adult human cardiomyocytes and generated force and action potentials consistent with normal fetal cardiomyocytes, documenting these parameters in hESC-derived cardiomyocytes and validating that the selected cells retained normal differentiation and function. CONCLUSION/SIGNIFICANCE: The protocols, vectors and gene expression data comprise tools to enhance cardiomyocyte production for large-scale applications.

Pubmed ID: 19352491


  • Kita-Matsuo H
  • Barcova M
  • Prigozhina N
  • Salomonis N
  • Wei K
  • Jacot JG
  • Nelson B
  • Spiering S
  • Haverslag R
  • Kim C
  • Talantova M
  • Bajpai R
  • Calzolari D
  • Terskikh A
  • McCulloch AD
  • Price JH
  • Conklin BR
  • Chen HS
  • Mercola M


PloS one

Publication Data

April 8, 2009

Associated Grants

  • Agency: NHLBI NIH HHS, Id: 5P01HL46345-12
  • Agency: NHGRI NIH HHS, Id: HG003053
  • Agency: NHLBI NIH HHS, Id: HL66621
  • Agency: NHLBI NIH HHS, Id: R33HL088266
  • Agency: NHLBI NIH HHS, Id: R37HL059502

Mesh Terms

  • Adult
  • Base Sequence
  • Cell Differentiation
  • DNA Primers
  • Drug Resistance
  • Embryonic Stem Cells
  • Fetal Proteins
  • Flow Cytometry
  • Gene Expression Profiling
  • Genetic Vectors
  • Green Fluorescent Proteins
  • Humans
  • Immunohistochemistry
  • Lentivirus
  • Myocardium
  • Promoter Regions, Genetic
  • Reverse Transcriptase Polymerase Chain Reaction
  • T-Box Domain Proteins