=========================================
*Spinacia oleracea* (spinach) cell lines
=========================================

Plant cell lines
================

* :doc:`/cell_lines/rpc00016`

* :doc:`/cell_lines/rpc00017`

* :doc:`/cell_lines/rpc00018`

Genotypes
=========

  .. csv-table::
     :header: "Cell line", "rbcL", "Marker 8", "NR intron"
     :widths: auto

     "\ :doc:`Spi-WT </genotyping/g_rpc00016>`\ ", "*Spinacia oleracea*", "Spi-WT/Spi-12F", "Spi-WT"
     "\ :doc:`Spi-I-1 </genotyping/g_rpc00017>`\ ", "*Spinacia oleracea*", "Spi-I-1", "Spi-I-1/Spi-12F"
     "\ :doc:`Spi-12F </genotyping/g_rpc00018>`\ ", "*Spinacia oleracea*", "Spi-WT/Spi-12F", "Spi-I-1/Spi-12F"

Protocols
=========

DNA barcoding --- rbcL
----------------------

  :doc:`_rbcL`

DNA sequencing
--------------

#. PCR

   * Primer set 《:doc:`_primers`》

       .. csv-table::
          :header: "Target", "Forward primer", "Reverse primer", "Product size"
          :widths: auto

          "Marker 8", "Spi8_F", "Spi8_R", "600 bp"
          "NR intron", "Sp_NR_F6", "Sp_NR_R1", "800 bp"

   * PCR reaction mixture

       .. csv-table::
          :header: "Component", "10 µL reaction"
          :widths: auto

          "TaKaRa Ex Taq™ Premix", "5 µL"
          "Forward primer (100 µM)", "0.125 µL"
          "Reverse primer (100 µM)", "0.125 µL"
          "Template DNA", "1 µL"
          "Sterile distilled water", "3.75 µL"

   * PCR reaction

       +-----------+------+---------+
       |Temperature|Time  |Cycles   |
       +===========+======+=========+
       |94°C       |1 min |         |
       +-----------+------+---------+
       |94°C       |30 sec|35 cycles|
       +-----------+------+         +
       |55°C       |30 sec|         |
       +-----------+------+         +
       |72°C       |1 min |         |
       +-----------+------+---------+
       |72°C       |7 min |         |
       +-----------+------+---------+

#. Check the DNA sequence of the PCR product.

.. 要確認： PCR reaction mixture、PCR reaction