The genomic DNA of potato plant leaf tissue extraction using CTAB method (Murray and Thompson, 1980) involves following Steps.

1. 100 mg of leaves from young leaves of potato plant are ground (in the liquid nitrogen) to a fine powder in the pre-chilled autoclaved pestle and mortar.
2. The powder is transferred to 1.5 ml eppendorf tube containing 750 μl pre-warmed extraction buffer (100mM Tris-Cl pH 8.0, 20 mM EDTA pH 8.0, 1.4 M NaCl, 2% CTAB and 0.2% 2-mercaptoethanol), mix by gentle inversion and incubate at 65^ºC for one hour.
3. After incubation, add 750 μl of chloroform: isoamylalcohol (24:1). Mix by gentle inversion and centrifuge at 12,000 rpm (Sigma) for 20 min at room temperature.
4. The upper aqueous phase is transferred to another tube without disturbing the inter-phase.
5. 2/3 volume of isopropanol is added to it and centrifuged at 12,000 rpm for 20 min at room temperature.
6. Supernatant is decanted and wash the pellet with 70% ethanol at 10,000 rpm for 5-10 min at 4^ºC.
7. The pellet is vacuum dried and dissolved in the 500 μl sterile water.
8. RNase is added to the dissolved pellet to a final concentration of 100 μg/ ml and incubate at 37 ^ºC for one hour.
9. Equal amount phenol: chloroform: Isoamylalcohol (25:24:1) is added and centrifuged at 12,000 rpm for 15 min.
10. Upper aqueous layer is carefully transferred to another tube and equal volume of chloroform: isoamylalcohol is added to it and mixed thoroughly by gentle inversion. This is centrifuged at 12,000 rpm for 15 min and upper layer is transferred to another eppendorf tube.
11. 1/10 volume of 3M sodium acetate (pH5.2) and 2 volumes of absolute alcohol are added to it and kept on ice for 20 minutes.This is centrifuged at 12,000 rpm for 10 min.
12. Wash the pellet with 70% alcohol. The pellet is vacuum dried, dissolved in 100 μl sterile water. The concentration of DNA sample is checked by spectrophotometer/ or NanoDrop and Qualitative estimation of genomic DNA is checked on 0.8% agarose gel using horizontal gel electrophoresis method.

• DNA extraction Protocol through standard genomic isolation kit:

  Total genomic DNA is extracted from fresh /frozen potato plant leaf tissue or tuber following manufacturer’s protocol (Gene Elute Plant Genomic Mini Prep Kit, Sigma Aldrich, U S A). Major Steps involved are as follows:

  1. Cells Disruption: Plant/tuber leaf tissue (about 200 mg) is ground into a fine powder in liquid nitrogen using a mortar and pestle. Transfer the powder to a microcentrifuge tube. Keep the sample on ice for immediate use.

  2. Cells Lyses: Add 350µl of Lysis solution [Part A] and 50µl of Lysis solution [Part B]. White precipitate is formed upon the addition of lysis solution. Incubate the mixture at 65°c for 10 minutes with occasional inversion top to dissolve the precipitate.

  3. Precipitation of Debris: Add 130µl of precipitation solution to the mixture, mix completely by inversion and place the sample on ice for 5 minutes. Centrifuge the sample at maximum speed (12,000-16000 ×g) for 5 minutes to pellet the cellular debris, protein and polysaccharides.

  4. Filtration of Debris: Carefully pipette the supernatant from step 3 onto a Gen Elute filtration column. Centrifuge at maximum speed (12,000-16000 ×g) for 1 minute. This removes any cellular debris not removed in step 3. Discard the filtration column, but retain the collection tube.

  5. Preparation for Binding: Add 700µl of binding solution directly to the flow- through liquid from step 4, mix thoroughly by inversion.

  6. Prepared Binding Column: Insert a gene Elute miniprep binding column into a provided micro-centrifuge tube, add 500µl of the column preparation solution to each miniprep column and centrifuge at 12,000g for 30 seconds to 1 minute. Discard the flow-through liquid.

  7. Load lysate: Carefully pipette 700µl of the mixture from step5 on to the column prepared in step 6 and centrifuge at maximum speed (12,000-16000 ×g) for 1 minute. Discard the flow-through liquid, retain the collection tube. Apply the remaining lysate from step 5 on to the binding column. Repeat the centrifuge as above and discard the flow-through liquid and collection tube.

  8. First Column Wash: Ethanol is added to the wash solution concentrate. Place the binding column from step 7 in to a fresh 2 ml collection tube and applied 500µl of the diluted wash solution to the column. Centrifuge at maximum speed (12,000-16000 ×g) for 1 minute. Discard the flow-through liquid, but retain the collection tube.

  9. Second Column Wash: Apply another 500µl of the diluted wash solution to the column and centrifuge at maximum speed (12,000-16000 ×g) for 3 minutes to dry the column. Discard the flow through liquid and column was again given one minute spin to dry the column completely.

  10. Elute DNA: Transfer the binding column to a fresh 2 ml collection tube. Apply 100µl of pre-warmed (65°C) elution solution to the column and centrifuge at maximum speed (12,000-16000 ×g). Discard the column. The elute contain pure genomic DNA. Store at -20°C for future use.