Cycle 3: 1X Cycle 4: 80X Increase setpoint temperature after cycle 2 by 0.5ºC Melt curve data collection and analysis enabled... PCR Quantification Spreadsheet Data for SYBR-490 We
Trang 1Phụ lục 6: Dung dịch ethanol 70%
- C2H5OH 70 %
Phụ lục 7: Potassium acetate 3 M, pH 5,2
- Potassium acetate 3 mol/l
- Hòa tan trong nước siêu sạch
- Hấp tiệt trùng
- Bảo quản ở nhiệt độ phòng
Phụ lục 8: Kết quả đo OD các quy trình ly trích
Kết quả thí nghiệm lần 1:
1 OD260nm/OD280nm 1.524 1.88 1.305 1.679
Nồng độ DNA (µg) 1.824 1.416 1.683 1.923
2 OD260nm/OD280nm 1.888 1.678 1.825 1.731
Nồng độ DNA (µg) 6.06 3.135 5.787 3.276
Nồng độ DNA (µg) 10.626 8.769 15.237 10.284
Kết quả thí nghiệm lần 2:
1 OD260nm/OD280nm 1.254 1.73 1.872 1.847
Nồng độ DNA (µg) 2.208 1.017 1.539 1.191
2 OD260nm/OD280nm 1.848 1.875 1.696 1.594
Nồng độ DNA (µg) 7.674 6.756 5.856 4.389
Nồng độ DNA (µg) 13.155 11.559 11.205 10.803
Trang 2Kết quả thí nghiệm lần 3:
1 OD260nm/OD280nm 1.815 1.509 1.667 1.864
Nồng độ DNA (µg) 1.503 2.286 2.691 1.73
2 OD260nm/OD280nm 1.245 1.208 1.755 1.697
Nồng độ DNA (µg) 4.134 5.022 3.375 5.0505
Nồng độ DNA (µg) 10.767 10.911 11.268 11.352
Trang 3Phụ lục 9: Kết quả phản ứng Real-Time PCR với thuốc nhuộm SYBR Green I
Đợt 1:
PCR Quantification with Melt Curve Report PCR Base Line Subtracted Curve Fit Data
Data generated on: 28-Jul-05 at 12:47 PM
Optical data file name: Data 28-Jul-05 1247-SYBR Green 35S-GMO.odm
Plate Setup file used: SYBR-Green 35S.psm
Protocol file used: SYBR Green-35S.tmo
Well factor collection: Experimental Plate
Comments
Protocol
Cycle 1: ( 1X)
Cycle 2: ( 50X)
Data collection and real-time analysis enabled
Cycle 3: ( 1X)
Cycle 4: ( 80X)
Increase setpoint temperature after cycle 2 by 0.5ºC
Melt curve data collection and analysis enabled
Cycle 5: ( 1X)
PCR Amp/Cycle Graph for SYBR-490
Trang 4Data Analysis Parameters
Calculated threshold using the maximum curvature approach is 20.0
Per-well baseline cycles have been determined automatically
Data analysis window is set at 95.00% of a cycle, centered at end of the cycle
Weighted Mean digital filtering has been applied Global filtering is off
PCR Quantification Spreadsheet Data for SYBR-490
Well Identifier Ct Setpoint
D02 Bt-176 1% 27.56
Melt Curve Graph for SYBR-490
Melt Curve Analysis Parameters
Weighted Mean digital filtering has been applied Global filtering is off
Threshold for automatic peak detection is set at 1.00
Melt Curve Analysis Spreadsheet Data for SYBR-490
Well Well Identifier Peak ID Melt Temp Beg Temp End Temp Peak Descriptor
Trang 5
Đợt 2:
PCR Quantification with Melt Curve Report PCR Base Line Subtracted Curve Fit Data
Data generated on: 05-Aug-05 at 03:35 PM
Optical data file name: Data 05-Aug-05 1505 SYBR Green -dot 5-3.odm
Plate Setup file used: SYBR Green 35S dot 5 5-8-0518.psm
Protocol file used: SYBR Green-35S-2.tmo
Well factor collection: Experimental Plate
Comments
Protocol
Cycle 1: ( 1X)
Cycle 2: ( 50X)
Data collection and real-time analysis enabled
Cycle 3: ( 1X)
Cycle 4: ( 80X)
Increase setpoint temperature after cycle 2 by 0.5ºC
Melt curve data collection and analysis enabled
Cycle 5: ( 1X)
PCR Amp/Cycle Graph for SYBR-490
Data Analysis Parameters
Trang 6Calculated threshold has been replaced by the user selected threshold 68.8
User selected baseline cycles are 2 to 10
Data analysis window is set at 95.00% of a cycle, centered at end of the cycle
Weighted Mean digital filtering has been applied Global filtering is off
PCR Quantification Spreadsheet Data for SYBR-490
Well Identifier Ct Setpoint
D02 B1 20.62
F07 B10 24.04
D06 B11 41.04
D04 B2 16.96
B04 B3 N/A
D10 B4 22.22
B06 B5 N/A
F03 B6 21.15
F05 B7 22.22
F09 B9 N/A
Melt Curve Graph for SYBR-490
Melt Curve Analysis Parameters
Weighted Mean digital filtering has been applied Global filtering is off
Threshold for automatic peak detection is set at 1.00
Melt Curve Analysis Spreadsheet Data for SYBR-490
Well Well Identifier Peak ID Melt Temp Beg Temp End Temp Peak Descriptor
Trang 7D4.3 81.5 78.5 82.0
D10 B4 D10.1 86.5 82.0 91.0
Trang 8
Đợt 3:
PCR Standard Curve Report with PCR Amp Cycle Graph PCR Base Line Subtracted Curve Fit Data
Comments
Protocol
Cycle 1: ( 1X)
Cycle 2: ( 50X)
Data collection and real-time analysis enabled
Cycle 3: ( 1X)
Cycle 4: ( 80X)
Increase setpoint temperature after cycle 2 by 0.5ºC
Melt curve data collection and analysis enabled
Cycle 5: ( 1X)
PCR Amp/Cycle Graph for SYBR-490
Trang 9Standard Curve Graph for SYBR-490
Data Analysis Parameters
Calculated threshold has been replaced by the user selected threshold 18.2
User selected baseline cycles are 2 to 24
Data analysis window is set at 95.00% of a cycle, centered at end of the cycle
Weighted Mean digital filtering has been applied Global filtering is off
Standard Curve Spreadsheet Data for SYBR-490 Units: copy number
Point
B02 Standard ST 35S 1 22.99 3.709 5.12E+03 5.12E+03 N/A 22.99 N/A
B03 Standard ST 35S 2 24.32 2.806 6.40E+02 6.40E+02 N/A 24.32 N/A
B04 Standard ST 35S 3 25.26 1.903 8.00E+01 8.00E+01 N/A 25.26 N/A
D02 Unknown Mau 1 copy 2 27.58 0.106 1.28E+00 8.88E+00 1.08E+01 26.88 9.86E-01
F02 Unknown Mau 1 copy 2 26.18 1.217 1.65E+01 8.88E+00 1.08E+01 26.88 9.86E-01
Melt Curve Graph for SYBR-490
Trang 10Melt Curve Analysis Parameters
Weighted Mean digital filtering has been applied Global filtering is off
Threshold for automatic peak detection is set at 1.00
Melt Curve Analysis Spreadsheet Data for SYBR-490
Well Well Identifier Peak ID Melt Temp Beg Temp End Temp Peak Descriptor
Trang 11Phụ lục 10: Kết quả chạy phản ứng Real-Time PCR với mẫu dò Taqman
Đợt 1: Đường chuẩn p35S
PCR Standard Curve Report with PCR Amp Cycle Graph PCR Base Line Subtracted Data
Data generated on: 22-Jul-05 at 06:15 PM
Optical data file name: Data 22-Jul-05 1815-Truong-35S-dot 1-sua2.odm
Plate Setup file used: GMO-35S.psm
Protocol file used: GMO-35S.tmo
Well factor collection: Experimental Plate
Comments
Protocol
Cycle 1: ( 1X)
Cycle 2: ( 45X)
Data collection and real-time analysis enabled
Cycle 3: ( 1X)
PCR Amp/Cycle Graph for FAM-490
Standard Curve Graph for FAM-490
Trang 12Data Analysis Parameters
Calculated threshold using the maximum correlation coefficient approach is 33.9
User selected baseline cycles are 2 to 20
Data analysis window is set at 95.00% of a cycle, centered at end of the cycle
Weighted Mean digital filtering has been applied Global filtering is off
Standard Curve Spreadsheet Data for FAM-490 Units: copy number
Point
E02 Standard Stan Corn-Ref 5 26.83 3.709 5.12E+03 5.12E+03 N/A 26.83 N/A
E04 Standard Stan Corn-Ref 6 30.04 2.806 6.40E+02 6.40E+02 N/A 30.04 N/A
E06 Standard Stan Corn-Ref 7 33.07 1.903 8.00E+01 8.00E+01 N/A 33.07 N/A
Đợt 2: Định lƣợng bắp chuyển gen Bt 176 1% (lần 1) Promoter 35S:
PCR Standard Curve Report with PCR Amp Cycle Graph PCR Base Line Subtracted Data
Data generated on: 22-Jul-05 at 06:15 PM
Optical data file name: Data 22-Jul-05 1815-Truong-35S-dot 1-sua2.odm
Plate Setup file used: GMO-35S.psm
Protocol file used: GMO-35S.tmo
Well factor collection: Experimental Plate
Comments
Protocol
Cycle 1: ( 1X)
Trang 13Step 1: 95.0ºC for 10:00
Cycle 2: ( 45X)
Data collection and real-time analysis enabled
Cycle 3: ( 1X)
PCR Amp/Cycle Graph for FAM-490
Standard Curve Graph for FAM-490
Data Analysis Parameters
Calculated threshold has been replaced by the user selected threshold 67.5
User selected baseline cycles are 2 to 20
Data analysis window is set at 95.00% of a cycle, centered at end of the cycle
Weighted Mean digital filtering has been applied Global filtering is off
Standard Curve Spreadsheet Data for FAM-490 Units: copy number
Point
E02 Standard Stan Corn-Ref 5 27.88 3.709 5.12E+03 5.12E+03 N/A 27.88 N/A
E04 Standard Stan Corn-Ref 6 31.35 2.806 6.40E+02 6.40E+02 N/A 31.35 N/A
E06 Standard Stan Corn-Ref 7 34.25 1.903 8.00E+01 8.00E+01 N/A 34.25 N/A
E08 Standard Stan Corn-Ref 8 36.22 1.000 1.00E+01 1.00E+01 N/A 36.22 N/A
E11 Unknown Bt 10 32.59 2.302 2.00E+02 2.00E+02 N/A 32.59 N/A