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Brochures & Specifications: TFA Aminolink™ Phosphoramidite 5' Labeling Reagent for DNA Synthesis: DNA Synthesis Reagents: Product Bulletin Product Literature

Photoreactive (caged) cyclic AMP analogs induce DNA synthesis in mammary epithelial cells. Citations & References

  • Authors: Yuh IS, Sheffield LG
  • Journal: Cell Biol Int
  • PubMed ID: 7550068
Catalog # D1037(Discontinued)

Membrane depolarization was required to induce DNA synthesis in murine macrophage cell line PU5-1.8. Citations & References

  • Authors: Kong SK, Suen YK, Choy YM, Fung KP, Lee CY
  • Journal: Immunopharmacol Immunotoxicol
  • PubMed ID: 1940052
Catalog #

Direct electrical detection of DNA synthesis. Citations & References

  • Authors: Pourmand N, Karhanek M, Persson HH, Webb CD, Lee TH, Zahradníková A, Davis RW
  • Journal: Proc Natl Acad Sci U S A
  • PubMed ID: 16614066
Catalog # 28100

Optimal lysophosphatidic acid-induced DNA synthesis and cell migration but not survival require intact autophosphorylation sites of the epidermal growth factor receptor. Citations & References

  • Authors: Deng W; Poppleton H; Yasuda S; Makarova N; Shinozuka Y; Wang DA; Johnson LR; Patel TB; Tigyi G
  • Journal: The Journal of Biological Chemistry
Catalog #

How are oligonucleotide primers synthesized? Product FAQ

Answer

Oligos are made using a DNA synthesizer, which is basically a computer-controlled reagent delivery system. The first base is attached to a solid support, usually a glass or polystyrene bead, which is designed to anchor the growing DNA chain in the reaction column. DNA synthesis consists of a series of chemical reactions.

(1) Deblocking
The first base, attached to the solid support via a chemical linker arm, is deprotected by removing the trityl protecting group. This produces a free 5' OH group to react with the next base.

(2) Coupling
The next base is added, which couples to the first base.

(3) Capping
Any of the first bases, which failed to react are capped. These failed bases will play no further part in the synthesis cycle.

(4) Oxidation
The bond between the first base and successfully coupled second base is oxidized to stabilize the growing chain.

(5) Deblocking
The 5' trityl group is removed from the base, which has been added.

Each cycle of reactions results in the addition of a single DNA base. A chain of DNA bases can be built by repeating the synthesis cycles until the desired length is achieved.

Answer Id:: E2931

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Parathyroid hormone depresses cytosolic pH and DNA synthesis in osteoblast-like cells. Citations & References

  • Authors: Reid IR, Civitelli R, Avioli LV, Hruska KA
  • Journal: Am J Physiol
  • PubMed ID: 2839040
Catalog #

A strategy for rapid cDNA cloning from double-stranded RNA templates isolated from plants infected with RNA viruses by using DNA polymerase Citations & References

  • Authors: Zhang YP, Rowhani A
  • Journal: J Virol Methods
  • PubMed ID: 10644087
Catalog # K200001

Synthesis and properties of the new thiol-specific reagent difluorescein disulfide: its application on histone-histone and histone-DNA interactions. Citations & References

  • Authors: Wingender E, Arellano A
  • Journal: Anal Biochem
  • PubMed ID: 6926748
Catalog # A458

What is included in the Ion AmpliSeq Kit for Chef DL8? Do I have to get any additional reagents? Product FAQ

Answer

The Ion AmpliSeq Kit for Chef DL8 enables robust, automated Ion AmpliSeq library preparation from genomic DNA or RNA using the Ion Chef Instrument and 1- or 2-pool Ion AmpliSeq primer panels. The kit contains sufficient reagents and consumables to prepare up to 32 barcoded and equalized libraries that are ready for template preparation using either the Ion Chef System or the Ion OneTouch 2 System. Library preparation from RNA additionally requires the SuperScript VILO cDNA Synthesis Kit, ordered separately. The kit components are as follows:

- Ion AmpliSeq Chef Reagents DL8 (4 cartridges)
- Ion AmpliSeq Chef Solutions DL8 (4 cartridges)
- Ion AmpliSeq Chef Supplies DL8 (1 box with 4 inserts)
Per insert:
-- Ion AmpliSeq Tip Cartridge DL8
-- Frames PCR Foil Seal
-- Enrichment Cartridge

- IonCode 0101-0132 in 96-well PCR Plates (dried) (1 set of 4 plates):
- IonCode 0101-0108 in 96 Well PCR Plate (red)
- IonCode 0109-0116 in 96 Well PCR Plate (yellow)
- IonCode 0117-0124 in 96 Well PCR Plate (green)
- IonCode 0125-0132 in 96 Well PCR Plate (blue)

Answer Id:: E11505

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DNA-binding transferrin conjugates as functional gene-delivery agents: synthesis by linkage of polylysine or ethidium homodimer to the transferrin carbohydrate moiety. Citations & References

  • Authors: Wagner E, Cotten M, Mechtler K, Kirlappos H, Birnstiel ML
  • Journal: Bioconjug Chem
  • PubMed ID: 1772904
Catalog # E1169

What kinds of cell health and viability assays can be performed by flow cytometry? Product FAQ

Answer

The following cell health and viability assays can be performed by flow cytometry :

-Apoptosis Assays:
Membrane Asymmetry: Annexin V is a member of a family of structurally related proteins that bind phospholipids in the presence of Ca2+. Annexin V binds several phospholipids, but shows highest affinity for phosphatidylserine.
Phosphatidylserine is normally found in the inner leaflet of the cell membrane; however, in the early stages of apoptosis, phosphatidylserine is observed to translocate to the outer leaflet. This translocation makes phosphatidylserine available for annexin V binding in the presence of Ca2+ containing incubation buffer. Cells undergoing apoptosis will stain with annexin V, while normal cells will not. annexin V is available conjugated with a wide range of fluorophores.

Mitochondrial Health: A distinctive feature of the early stages of apoptosis is the disruption of the mitochondria, including changes in membrane and redox potential. We exclusively offer a number of fluorescent probes for analyzing mitochondrial activity in live cells by flow cytometry, with minimal disruption of cellular function.

The MitoProbe family of mitochondrial stains (MitoProbe DiOC2(3) Assay Kit, Cat. No. M34150, MitoProbe JC-1 Assay Kit, Cat. No. M34152, and MitoProbe DiIC1(5) Assay Kit, Cat. No. M34151) provides quick, easy, and reliable flow cytometric detection of the loss of mitochondrial membrane potential that occurs during apoptosis.

Caspase Activity: The CellEvent Caspase-3/7 Green Flow Cytometry Assay Kit (Cat. No. C10427) enables flow cytometric detection of activated caspase-3 and caspase-7 in apoptotic cells. The kit includes the novel fluorogenic substrate CellEvent Caspase-3/7 Green Detection Reagent which targets the recognition sequence for activated caspase-3 and caspase-7, as well as SYTOX AADvanced Dead Cell Stain.

DNA Fragmentation: The later stages of apoptosis are characterized by changes in nuclear morphology, including DNA fragmentation, chromatin condensation, degradation of nuclear envelope, nuclear blebbing, and DNA strand breaks. DNA fragmentation that occurs during apoptosis produces DNA strand breaks, and can be analyzed using TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) assays. The APO-BrdU TUNEL assay (Cat. No. A23210) is a two-color assay for labeling DNA breaks and total cellular DNA to detect apoptotic cells by imaging or flow cytometry.

Nuclear Chromatin Condensation: The later stages of apoptosis are characterized by changes in nuclear morphology, including DNA fragmentation, chromatin condensation, degradation of nuclear envelope, nuclear blebbing, and DNA strand breaks. Cells undergoing apoptosis display an increase in nuclear chromatin condensation. As the chromatin condenses, cell-permeable nucleic acid stains becomes hyperfluorescent, thus enabling the identification of apoptotic cells when combined with a traditional dead-cell stain. The Vybrant Apoptosis Assay Kit #5, Hoechst 33342/Propidium Iodide (Cat. No. V13244) provides a rapid and convenient assay for apoptosis based on fluorescence detection of the compacted state of the chromatin in apoptotic cells. The Chromatin Condensation & Membrane Permeability Dead Cell Apoptosis Kit with Hoechst 33342, YO-PRO-1, and PI dyes, for flow cytometry (Cat. No. V23201) detects apoptotic cells with changes in nuclear chromatin condensation and plasma membrane permeability.

-Cell Cycle Analysis:
Live cell assays: The Vybrant DyeCycle family of dyes offers robust fluorescent dyes for live-cell cycle analysis with limited cytotoxicity using 405 nm (Cat. No. V35003), 488 nm (Cat. No. V35004), 532 nm (Cat. No. V35005), or 633 nm (Cat. Nos. V10309 and V10273) excitation. The dyes have low cytotoxicity, allowing stained cells to be sorted and otherwise cultured or assessed with functional assays after staining.

Fixed cell assays: Analyzing cell cycle using FxCycle Violet Stain (Cat. No. F10347), SYTOX AADvanced Dead Cell Stain Kit (Cat. No. S10349) or FxCycle Far Red Stain (Cat. No. F10348) allows for multiple color options for simplified fixed cell cycle analysis.

-Cell Proliferation:
Dye dilution assays for cell proliferation: Dye dilution assays for cell proliferation rely on cell membrane–permeant fluorescent molecules. Upon entry into the cell, the dye will covalently bind to amine groups on proteins, resulting in long-term dye retention within the cell. Through subsequent cell divisions, each daughter cell receives approximately half the fluorescence of the parent. Analysis of the fluorescence intensities of cell populations by flow cytometry enables determination of the number of generations through which a cell or population has progressed since the label was applied. CellTrace fluorescent stains can be used without affecting morphology or physiology to trace generations in vivo or in vitro. There is no known effect on proliferative ability or biology of cells and they are well retained in cells for several days post-stain. Available kits for flow cytometry include CellTrace CFSE Cell Proliferation Kit (Cat. No. C34554), CellTrace Violet Cell Proliferation Kit (Cat. No. C34557), and CellTrace Far Red Cell Proliferation Kit (Cat. No. C34564).

DNA Synthesis Assays: Measuring the synthesis of new DNA is a precise way to assay cell proliferation in individual cells or in cell populations. DNA synthesis–based cell proliferation assays measure the rate of new DNA synthesis based on incorporation of modified nucleosides. The Click-iT Plus EdU cell proliferation assay utilizes the power of click chemistry and the modified nucleoside EdU to provide a superior alternative to BrdU staining for detecting and quantitating newly synthesized DNA. The Click-iT Plus EdU cell proliferation assay is available with Pacific Blue (Cat. No. C10636), Alexa Fluor 488 (Cat. Nos. C10632 and C10633), and Alexa Fluor 647 (Cat. Nos. C10634 and C10635).

-Viability Assays:
Dead cells often give false positive results, as they tend to bind non-specifically to many reagents. Therefore, removing dead cells from your flow cytometry data is a critical step to help ensure accurate results and analysis.

Non-fixable Membrane Permeability Stains: SYTOX Dead Cell Stains (Cat. Nos. S34857, S34860, S34861, S34859, and S34862) do not cross intact cell membranes, and they exhibit increased fluorescence upon dsDNA binding, making them some of our most brilliant dead cell stains. Cell-impermeant classic DNA-binding dyes include propidium iodide (Cat. No. P21493) and 7-AAD (Cat. No. A1310). Both of these dyes have been used extensively for viability assays in flow cytometry. CellTrace Calcein AM dyes can be passively loaded into adherent and nonadherent cells. These cell-permeant esterase substrates serve as viability probes that measure both enzymatic activity, which is required to activate their fluorescence, and cell membrane integrity, which is required for intracellular retention of their fluorescent products. Available with blue (Cat. No. C34853), violet (Cat. No. C34858), and green (Cat. No. C34852) fluorescence, these dyes are ideal for short-term staining of live cells and can be used in multiplexed flow cytometry experiments.

Fixable Viability Stains: The LIVE/DEAD Fixable Dead Cell Stains are fixable viability dyes that help to ensure accurate assessment of cell viability in samples after fixation and/or permeabilization. LIVE/DEAD Fixable Dead Cell Stain Kits are based on the reaction of a fluorescent reactive dye with cellular proteins (amines). These dyes cannot penetrate live-cell membranes, so only cell-surface proteins are available to react with the dye, resulting in dim staining. The reactive dye can permeate the damaged membranes of dead cells and stain both the interior and exterior amines, resulting in more intense staining. LIVE/DEAD Fixable Dead Cell Stain Kits are available in eight single-channel colors available for UV, 405, 488, 532, 561, or 633 nm lasers in three packaging sizes to match your experim

Answer Id:: E14826

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I'm having a hard time amplifying/cloning low copy cDNAs via PCR. Do you have any suggestions for me? Product FAQ

Answer

You may want to consider using our Platinum SuperFi II DNA Polymerase (Cat. No. 12361010) and try optimizing your PCR conditions if needed. The Platinum SuperFi II DNA Polymerase demonstrates a superior detection sensitivity with as little as 0.4 ng of genomic DNA. It is also a good idea to include a positive control reaction in parallel to your test PCR to ensure there are no issues with your reagents during handling or use over time. Please note that control templates may be ordered through our GeneArt Gene Synthesis (de novo sequence with 100% guarantee delivered in a vector) or Strings (up to 3 Kb, blunt-ended dsDNA pools, cloned into ZeroBlunt TOPO vector, convenient for multiple controls) services.

Answer Id:: E18628

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How many samples can I process with the Oncomine Focus Assay, Chef-Ready Library? Product FAQ

Answer

The Oncomine Focus Assay, Chef-Ready Library contains the following components needed to prepare libraries for 32 samples:

- Oncomine Focus Assay, DNA Chef-Ready Panel (2X) - contains the DNA primer pool
- Oncomine Focus Assay, RNA Chef-Ready Panel (2X) - contains the RNA primer pool
- Ion AmpliSeq Kit for Chef Reagents DL8 cartridge - allows automation of the library prep on the Ion Chef Instrument

Note: The SuperScript transcriptase for cDNA synthesis is not included and must be purchased separately.

Answer Id:: E17245

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