TCH CHIT DNA BNG BUFFER LY GII 1. Add 500 ul Lysis Buffer in a 1.5ml tube. 2.

Put shrimp sample into the tube and grind it with a disposable grinder. 3. Incubate the prepared sample at 95 for 10 minutes, then centrifuge at 12000g (12000 rpm r=5~7cm) for 10 minutes. 4. Transfer 200 ul of the upper clear solution to a fresh 1.5ml tube with 400 ul 95% ethanol. 5. Vortex briefly, centrifuge at 12000g for 5 minutes, then decant the ethanol and dry the pellet. 6. Dissolve the pellet by ddH2O or TE buffer. 7. If sample needed to be preserved for longer period, TE buffer is recommended. Sample can be stored in -20 for one year. 8. Please fine tune the volume of ddH2O or TE buffer according to the real recovery efficiency. TCH CHIT DNA BNG DTAB_CTAB 1. Place about 20 mg specimen into a 2ml tube containing 0.6 ml DTAB solution. 2. Grind the sample in the tube with a disposable grinder. 3. Incubate the prepared sample at 75 for 5 minutes, then cool down to room temperature. 4. Vortex briefly and spin down the mixture, then add 0.7 ml of chloroform, vortex for another 20 seconds and centrifuge at 12000g (12000 rpm, r = 5~7 cm) for 5 minutes. 5. Transfer 200 ul of the upper aqueous phase to a new 1.5ml tube. Add 100 ul of CTAB Solution and 900 ul ddH2O. Vortex briefly, then incubate at 75 for 5 minutes. 6. Cool down to room temperature and centrifuge at 12000g for 10 minutes. 7. Carefully decant the supernatant, resuspend the pellet with 150 ul Dissolving Solution, incubate at 75 for 5 minutes then cool down to room temperature. 8. Spin at 12000g for 5 minutes. Transfer the clear solution to a fresh 1.5ml tube with 300 ul of 95% ethanol. 9. Vortex briefly, centrifuge at 12000g for 5 minutes, then wash the pellet with 200 ul of 75% ethanol, spin down, dry the pellet and dissolve in ddH2O or TE buffer. 10. If sample needed to be preserved for longer period, TE buffer is recommended. Sample can be stored in -20 for one year. 11. Please fine tune the volume of ddH2O or TE buffer according to the real recovery efficiency.

TCH CHIT DNA BNG INSTAGENE MATRIX 1. Cut a small (3 mm3) piece of tissue on a fresh glass slide using a fresh scalpel blade. Macerate slightly with the blade and place in a sterile 1.5 ml screw cap Eppendorf tube. 2. Add 1 ml of 1x Trypsin/EDTA (Gibco BRL). Incubate at room temperature for 15-30 minutes, shaking occasionally. 3. Spin down (13K, 2-3 minutes) to pellet tissue, and carefully remove all the supernatant. 4. Add 250 l of InstaGene matrix (Bio-Rad Laboratories), and mix well. Incubate at 56 C for 30 minutes, mixing occasionally. 5. Vortex tube thoroughly for 10 seconds, place in boiling water bath or 100 C heat block for 8 minutes. 6. Vortex thoroughly for 10 seconds, spin at 13K for 2-3 minutes. Use 20 l of the supernatant for each PCR reaction. Notes: Store remainder at -20 C for subsequent PCRs. Repeat step 6 when reusing the preparation (allow to thaw first).