Laboratory Medicine ›› 2020, Vol. 35 ›› Issue (4): 354-362.DOI: 10.3969/j.issn.1673-8640.2020.04.016
• Orginal Article • Previous Articles Next Articles
DAI Qian1, HUANG Fei1, WANG Yupeng2, HUANG Ao2, CHENG Jianwen2, PAN Baishen1, GUO Wei1, ZHOU Jian2, FAN Jia2, YANG Xinrong2, WANG Beili1()
Received:
2019-09-23
Online:
2020-04-30
Published:
2020-05-19
CLC Number:
DAI Qian, HUANG Fei, WANG Yupeng, HUANG Ao, CHENG Jianwen, PAN Baishen, GUO Wei, ZHOU Jian, FAN Jia, YANG Xinrong, WANG Beili. Establishment of a digital PCR mutation detection platform for ctDNA in patients with intrahepatic cholangiocarcinoma and its clinical application[J]. Laboratory Medicine, 2020, 35(4): 354-362.
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URL: https://www.shjyyx.com/EN/10.3969/j.issn.1673-8640.2020.04.016
基因位点 | 引物序列(5'~3') | 引物大小/bp |
---|---|---|
KRAS G12D | ||
F | CCGAGTGGAAGGAAATTTGC | 20 |
R | CACCACCACACTATGTCGAAAAG | 23 |
Probe1 | TATTTGGATGACAAAAAC | 18 |
Probe2 | TGGATGACAGAAAC | 14 |
TP53 C242S | ||
F | CCTGAGGTTGGCTCTGACTGTA | 22 |
R | CCAGTGTGATGATGGTGAGGAT | 22 |
Probe1 | CATGAACGGGAGGC | 14 |
Probe2 | CATGAACCGGAGGC | 14 |
IDH1 R132C | ||
F | CTTGTGAGTGGATGGGTAAAACCTA | 25 |
R | CACATTATTGCCAACATGACTTACTTGAT | 29 |
Probe1 | AAGCATGACAACCTATG | 17 |
Probe2 | ATCATAGGTCATCATGC | 17 |
基因位点 | 引物序列(5'~3') | 引物大小/bp |
---|---|---|
KRAS G12D | ||
F | CCGAGTGGAAGGAAATTTGC | 20 |
R | CACCACCACACTATGTCGAAAAG | 23 |
Probe1 | TATTTGGATGACAAAAAC | 18 |
Probe2 | TGGATGACAGAAAC | 14 |
TP53 C242S | ||
F | CCTGAGGTTGGCTCTGACTGTA | 22 |
R | CCAGTGTGATGATGGTGAGGAT | 22 |
Probe1 | CATGAACGGGAGGC | 14 |
Probe2 | CATGAACCGGAGGC | 14 |
IDH1 R132C | ||
F | CTTGTGAGTGGATGGGTAAAACCTA | 25 |
R | CACATTATTGCCAACATGACTTACTTGAT | 29 |
Probe1 | AAGCATGACAACCTATG | 17 |
Probe2 | ATCATAGGTCATCATGC | 17 |
突变位点 | 靶值 | 第1次检测 | 第2次检测 | 第3次检测 | 均值 | 偏差 |
---|---|---|---|---|---|---|
KRASG12D | 5.00 | 4.60 | 4.40 | 4.30 | 4.43 | -11.40 |
1.00 | 1.21 | 0.90 | 1.09 | 1.07 | 7.00 | |
0.50 | 0.45 | 0.42 | 0.41 | 0.43 | -14.00 | |
TP53 C242S | 5.00 | 4.78 | 5.10 | 4.30 | 4.73 | -5.40 |
1.00 | 0.89 | 0.80 | 0.94 | 0.88 | -12.00 | |
0.50 | 0.43 | 0.36 | 0.49 | 0.43 | -14.00 | |
IDH1 R132C | 7.80 | 9.00 | 8.00 | 6.20 | 7.73 | -0.90 |
1.56 | 2.10 | 1.10 | 1.30 | 1.50 | -4.00 | |
0.78 | 0.82 | 1.01 | 0.75 | 0.86 | 10.26 |
突变位点 | 靶值 | 第1次检测 | 第2次检测 | 第3次检测 | 均值 | 偏差 |
---|---|---|---|---|---|---|
KRASG12D | 5.00 | 4.60 | 4.40 | 4.30 | 4.43 | -11.40 |
1.00 | 1.21 | 0.90 | 1.09 | 1.07 | 7.00 | |
0.50 | 0.45 | 0.42 | 0.41 | 0.43 | -14.00 | |
TP53 C242S | 5.00 | 4.78 | 5.10 | 4.30 | 4.73 | -5.40 |
1.00 | 0.89 | 0.80 | 0.94 | 0.88 | -12.00 | |
0.50 | 0.43 | 0.36 | 0.49 | 0.43 | -14.00 | |
IDH1 R132C | 7.80 | 9.00 | 8.00 | 6.20 | 7.73 | -0.90 |
1.56 | 2.10 | 1.10 | 1.30 | 1.50 | -4.00 | |
0.78 | 0.82 | 1.01 | 0.75 | 0.86 | 10.26 |
项目 | 批内精密度 | 批间精密度 | |||||
---|---|---|---|---|---|---|---|
x | s | CV/% | x | s | CV/% | ||
KRASG12D | |||||||
高丰度 | 21.78 | 1.20 | 5.50 | 21.82 | 1.13 | 5.19 | |
低丰度 | 2.47 | 0.28 | 11.46 | 2.47 | 0.28 | 11.46 | |
TP53 C242S | |||||||
高丰度 | 19.68 | 0.85 | 4.32 | 19.03 | 0.83 | 4.36 | |
低丰度 | 2.21 | 0.17 | 7.51 | 2.37 | 0.35 | 14.57 | |
IDH1 R132C | |||||||
高丰度 | 16.49 | 0.92 | 5.61 | 16.65 | 1.28 | 7.68 | |
低丰度 | 1.86 | 0.17 | 8.99 | 1.80 | 0.21 | 11.49 |
项目 | 批内精密度 | 批间精密度 | |||||
---|---|---|---|---|---|---|---|
x | s | CV/% | x | s | CV/% | ||
KRASG12D | |||||||
高丰度 | 21.78 | 1.20 | 5.50 | 21.82 | 1.13 | 5.19 | |
低丰度 | 2.47 | 0.28 | 11.46 | 2.47 | 0.28 | 11.46 | |
TP53 C242S | |||||||
高丰度 | 19.68 | 0.85 | 4.32 | 19.03 | 0.83 | 4.36 | |
低丰度 | 2.21 | 0.17 | 7.51 | 2.37 | 0.35 | 14.57 | |
IDH1 R132C | |||||||
高丰度 | 16.49 | 0.92 | 5.61 | 16.65 | 1.28 | 7.68 | |
低丰度 | 1.86 | 0.17 | 8.99 | 1.80 | 0.21 | 11.49 |
突变位点 | 预期值 | 实测值 | s | CV |
---|---|---|---|---|
KRASG12D | 0.001 | 0.010 | 0.004 2 | 42.30 |
0.01 | 0.008 0 | 0.003 1 | 38.75 | |
0.1 | 0.13 | 0.018 | 14.08 | |
1 | 0.82 | 0.043 | 5.35 | |
10 | 13 | 0.81 | 6.25 | |
100 | 114 | 7.44 | 6.52 | |
TP53 C242S | 0.001 | 0.008 0 | 0.003 3 | 40.63 |
0.01 | 0.030 | 0.007 9 | 26.33 | |
0.1 | 0.20 | 0.029 | 14.50 | |
1 | 1.12 | 0.043 | 3.89 | |
10 | 8 | 0.81 | 10.15 | |
100 | 106 | 6.54 | 6.17 | |
IDH1 R132C | 0.001 | 0.006 0 | 0.004 1 | 68.67 |
0.01 | 0.030 | 0.009 3 | 31.00 | |
0.1 | 0.20 | 0.037 | 18.50 | |
1 | 0.80 | 0.051 | 6.40 | |
10 | 11 | 0.71 | 6.47 | |
100 | 122 | 5.48 | 4.49 |
突变位点 | 预期值 | 实测值 | s | CV |
---|---|---|---|---|
KRASG12D | 0.001 | 0.010 | 0.004 2 | 42.30 |
0.01 | 0.008 0 | 0.003 1 | 38.75 | |
0.1 | 0.13 | 0.018 | 14.08 | |
1 | 0.82 | 0.043 | 5.35 | |
10 | 13 | 0.81 | 6.25 | |
100 | 114 | 7.44 | 6.52 | |
TP53 C242S | 0.001 | 0.008 0 | 0.003 3 | 40.63 |
0.01 | 0.030 | 0.007 9 | 26.33 | |
0.1 | 0.20 | 0.029 | 14.50 | |
1 | 1.12 | 0.043 | 3.89 | |
10 | 8 | 0.81 | 10.15 | |
100 | 106 | 6.54 | 6.17 | |
IDH1 R132C | 0.001 | 0.006 0 | 0.004 1 | 68.67 |
0.01 | 0.030 | 0.009 3 | 31.00 | |
0.1 | 0.20 | 0.037 | 18.50 | |
1 | 0.80 | 0.051 | 6.40 | |
10 | 11 | 0.71 | 6.47 | |
100 | 122 | 5.48 | 4.49 |
患者编号 | 突变位点 | 突变丰度(%) |
---|---|---|
3 | KRAS G12D | 0.25 |
7 | KRAS G12D | 0.12 |
TP53 C242S | 0.32 | |
9 | IDH1 R132C | 1.62 |
10 | KRAS G12D | 0.25 |
14 | TP53 C242S | 0.16 |
20 | KRAS G12D | 0.46 |
21 | IDH1 R132C | 1.32 |
患者编号 | 突变位点 | 突变丰度(%) |
---|---|---|
3 | KRAS G12D | 0.25 |
7 | KRAS G12D | 0.12 |
TP53 C242S | 0.32 | |
9 | IDH1 R132C | 1.62 |
10 | KRAS G12D | 0.25 |
14 | TP53 C242S | 0.16 |
20 | KRAS G12D | 0.46 |
21 | IDH1 R132C | 1.32 |
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