Page 37 - CASA Bulletin of Anesthesiology 2022, Vol 9, No 1 (1)
P. 37

Vol. 9, No 1, 2022



                                           表 1. 50 个恶性高热易感基因 SNP 位点


                     Panel  A          Panel  B         Panel  C          Panel  D         Panel  E


                      RYR1 c.487 C>T     RYR1 c.742      RYR1 c.103 T>C   RYR1 c.488 G>T      RYR1 c.1201 C>T
                                         G>A/C


                       RYR1 c.1021       RYR1 c.6488    RYR1 c.1209 C>G   RYR1 c.1589 G>A     RYR1 c.7063 C>T
                       G>A/C             G>A

                     RYR1 c.1840 C>T     RYR1 c.7048    RYR1 c.1565 A>C   RYR1 c.7360 C>T    RYR1 c.7042_7044 del
                                         G>A                                                    (GAG)

                     RYR1 c.6502 G>A     RYR1 c.7282    RYR1 c.6487 C>T    RYR1 c.14497       RYR1 c.7354 C>T
                                         G>A                                C>T


                     RYR1 c.6617 C>G/T   RYR1 c.7361    RYR1 c.7124 G>C   RYR1 c.14693 T>C    RYR1 c.11969 G>T
                                         G>A

                     RYR1 c.7300 G>A    RYR1 c.14477    RYR1 c.7372 C>T    RYR1 c.7522        RYR1 c.14545 G>A
                                         C>T                               C>T/G

                     RYR1 c.7373 G>A     RYR1 c.7007      RYR1 c.7523     RYR1 c.130 C>T      RYR1 c.7304 G>A
                                         G>A              G>A


                     RYR1 c.14582 G>A   RYR1 c.1654 C>T   RYR1 c.14512    RYR1 c.982 C>T      RYR1 c.1598 G>A
                                                           C>G

                      CACNA1S c.520                      RYR1 c.1841G>T
                       C>T

                     CACNA1S c.3257                         RYR1
                                                        c.14387A>G


                     RYR1 c.1597 C>T




                   2.3  多重 PCR

                   多重 PCR 体系包括 6 个组分:  MH PCR Master Mix、MH PCR Buffer、MH Primer Mix

               A/B/C/D/E、 MH 阴性对照、Size 500 Plus。其中，MH PCR Master Mix 由缓冲液、d NTP、
               pc DNA 质粒、HS Taq 聚合酶、BSA、甜菜碱等组成。具体实验操作时，按每反应配制下述体
               系：将 5μL MH PCR Master Mix、2μl MH Primer buffer、1μL MH Primer Mix
               A/B/C/D/E、1μL 模板、加水配制成 10μL PCR 体系。混匀后采用 Veriti 96-well 热循环仪

               (Applied Biosystems, California, USA) ，按下列循环标准孵育上述 PCR 反应混合物:  95℃ 5 分
               钟；94℃ 10 秒; 94℃30 秒，61℃ 1 分钟，70℃ 1 分钟，29 个循环；60℃处理 15 分钟，4℃保
               存。

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