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DNA modification of live cell surface.

Borisenko GG, Zaitseva MA, Chuvilin AN, Pozmogova GE - Nucleic Acids Res. (2009)

Bottom Line: By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface.All procedures can be completed within minutes and do not alter cell viability.Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Physical-Chemical Medicine, Russian Academy of Science, Moscow 119312, Russia. grigoryb@yahoo.com

ABSTRACT
We report a novel approach for the attachment of DNA fragments to the surface of live cells. By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface. Method permits to control amount of immobilized DNA on the cell surface. All procedures can be completed within minutes and do not alter cell viability. Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion. Thus, this method allows rapid and simple DNA multicoding of the cell surface and, therefore, opens new opportunities in manipulating with cell-cell interactions.

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MALDI TOF mass spectra of oligonucleotide conjugates. Peaks at m/z 6159, 6399 and 6427 represent FT18N, FT18Pal and FT18Ste respectively.
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Figure 3: MALDI TOF mass spectra of oligonucleotide conjugates. Peaks at m/z 6159, 6399 and 6427 represent FT18N, FT18Pal and FT18Ste respectively.

Mentions: Relatively short Т18 and Т25 homosequences were prepared to avoid any steric hindrance during hybridization near the cell surface. Oligomers were labeled with fluorescent moiety (6)-FAM to visualize both binding of conjugates to PM and hybridization of DNA at the cell surface. Reaction products were analyzed by using HPLC, MALDI-TOF mass spectrometry, UV spectroscopy and fluorescence spectroscopy. A typical mass spectra of 5′-(6)-FAM(T)18-OP(O)CH2-CH(CH2OH)-(CH2)4NH2 (FT18N), 5′-(6)-FAM(T)18-alkylaminostearate (FT18NSte) and 5′-(6)-FAM(T)18-alkylaminopalmitate (FT18NPal) are presented at Figure 3. Molecular masses of products obtained by MALDI-TOF MS deviated by less than 0.3% from anticipated estimates.Figure 3.


DNA modification of live cell surface.

Borisenko GG, Zaitseva MA, Chuvilin AN, Pozmogova GE - Nucleic Acids Res. (2009)

MALDI TOF mass spectra of oligonucleotide conjugates. Peaks at m/z 6159, 6399 and 6427 represent FT18N, FT18Pal and FT18Ste respectively.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651772&req=5

Figure 3: MALDI TOF mass spectra of oligonucleotide conjugates. Peaks at m/z 6159, 6399 and 6427 represent FT18N, FT18Pal and FT18Ste respectively.
Mentions: Relatively short Т18 and Т25 homosequences were prepared to avoid any steric hindrance during hybridization near the cell surface. Oligomers were labeled with fluorescent moiety (6)-FAM to visualize both binding of conjugates to PM and hybridization of DNA at the cell surface. Reaction products were analyzed by using HPLC, MALDI-TOF mass spectrometry, UV spectroscopy and fluorescence spectroscopy. A typical mass spectra of 5′-(6)-FAM(T)18-OP(O)CH2-CH(CH2OH)-(CH2)4NH2 (FT18N), 5′-(6)-FAM(T)18-alkylaminostearate (FT18NSte) and 5′-(6)-FAM(T)18-alkylaminopalmitate (FT18NPal) are presented at Figure 3. Molecular masses of products obtained by MALDI-TOF MS deviated by less than 0.3% from anticipated estimates.Figure 3.

Bottom Line: By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface.All procedures can be completed within minutes and do not alter cell viability.Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion.

View Article: PubMed Central - PubMed

Affiliation: Research Institute of Physical-Chemical Medicine, Russian Academy of Science, Moscow 119312, Russia. grigoryb@yahoo.com

ABSTRACT
We report a novel approach for the attachment of DNA fragments to the surface of live cells. By using fluorescence microscopy and flow cytometry we demonstrated that our synthetic conjugates of fatty acid with oligonucleotides can be incorporated in plasma membrane and then hybridized with complementary sequences at the cell surface. Method permits to control amount of immobilized DNA on the cell surface. All procedures can be completed within minutes and do not alter cell viability. Using this approach we tethered floating myeloid HL-60 cells to adherent A431 epitheliocytes in a sequence specific fashion. Thus, this method allows rapid and simple DNA multicoding of the cell surface and, therefore, opens new opportunities in manipulating with cell-cell interactions.

Show MeSH