Esophageal Adenocarcinoma Methods and Protocols

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is distributed equally across the slide. Take care to not touch
the slide!


  1. Avoid dropping the array slide onto the gasket slide as you may
    cause mixing between gasket wells.

  2. Recommended controls for this reaction should include a well
    where water replaces RNA in the reverse transcription reaction
    and wells with no reverse transcriptase as negative control.
    However, you can adjust your controls according to your
    experiment design and aims.

  3. Prior to opening the reverse transcription primer tubes, you
    should thaw them on ice. Subsequently, mix and centrifuge
    them.

  4. In the negative reverse transcriptase batch, replace the reverse
    transcriptase volume with RNase-free water.

  5. Reverse transcription master mix should be kept on ice while
    you prepare your microRNA samples.

  6. Adjust these volumes for the appropriate number of reverse
    transcription reactions and prepare separate batches for experi-
    mental miRNA and reference miRNA.

  7. You can use the amplification plots of the reference miRNA
    reaction to ensure that the assay is performing correctly.


References



  1. Ambros V (2004) The functions of animal
    microRNAs. Nature 431:350–355. https://
    doi.org/10.1038/nature02871

  2. Mamoori A, Gopalan V, Smith RA, Lam AK
    (2016) Modulatory roles of microRNAs in the
    regulation of different signalling pathways in
    large bowel cancer stem cells. Biol Cell 108:51–
    64. https://doi.org/10.1111/boc.201500062

  3. Salajegheh A, Vosgha H, Rahman MA, Amin
    M, Smith RA, Lam AK (2016) Interactive role
    of miR-126 on VEGF-A and progression of
    papillary and undifferentiated thyroid carci-
    noma. Hum Pathol 51:75–85. https://doi.
    org/10.1016/j.humpath.2015.12.018

  4. Gopalan V, Islam F, Pillai S, Tang JC, Tong
    DK, Law S, Chan KW, Lam AK (2016)
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    geal squamous cell carcinoma. Exp Cell Res
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  5. Gopalan V, Smith RA, Lam AK (2015)
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    tal cancers and its cellular effects. Exp Cell Res
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    yexcr.2014.08.006
    6. Chruscik A, Lam AK (2015) Clinical patho-
    logical impacts of microRNAs in papillary thy-
    roid carcinoma: a crucial review. Exp Mol
    Pathol 99:393–398. https://doi.
    org/10.1016/j.yexmp.2015.08.013
    7. Salajegheh A, Vosgha H, Md Rahman A, Amin
    M, Smith RA, Lam AK (2015) Modulatory role
    of miR-205 in angiogenesis and progression of
    thyroid cancer. J Mol Endocrinol 55:183–196.
    https://doi.org/10.1530/jme-15-0182
    8. Ebrahimi F, Gopalan V, Wahab R, Lu CT,
    Smith RA, Lam AK (2015) Deregulation of
    miR-126 expression in colorectal cancer patho-
    genesis and its clinical significance. Exp Cell
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    j.yexcr.2015.10.004
    9. Gopalan V, Pillai S, Ebrahimi F, Salajegheh A,
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Moein Amin et al.
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