应用实时荧光定量RT-PCR法建立肝癌分子诊断指数

表1 9个基因的引物序列

引物	序列5'→3'片段	大小(bp)	退火温度( ℃)
G3PDH	CTC ATG ACC ACA GTC CAT GCC ATC ACT G	140	55
	CAT GAG GTC CAC CAC CCT GTT GCT GTA
GPC3	TGA CCT GAA AGT ATT TGG GAA TTT CC	427	60
	CAA TAG TGG TGG TCA GCT TTC CTG
E2F1	GAA CCA CAT CCA GTG GCT GG	520	63
	GGA CAA CAG CGG TTC TTG CT
MMP2	GAA GGC TGT GTT CTT TGC AGG	325	61
	TCA GGT AAT AGG CAC CCT TGA AG
CLDN10	TGT CAG GGC TGT GCT CAA TG	150	57
	CCA CCA ATT ATG CAC AGT GAG G
HGF	GAA AAC AAT GCC TCT GGT TCC	200	57
	TGT GTT CGT GTG GTA TCA TGG A
PTEN	TAG AGT TCT TCC ACA AAC AGA ACA GA	550	60
	AAT CTG ACA CAA TGT CCT ATT GCC
DLC1	TTT ACC ACC TGC TGA ATT ACT CAC GCC	300	60
	ACG TCT GTG TAG TGC ACA GGA AAG CC
PRDM2	GGG ATG GAT GTG CAT TGA TG	350	61
	GCA GAG GTG AAA TCT GGC TCA
MGMT	GTG CAC AGC CTG GCT GAA T	500	62
	CGC ATC CGA TGC AGT GTT AC

表2 基因表达量和分子诊断指数

	GPC3	E2F1	MMP2	CLDN10	HGF	PTEN	DLC1	PRDM2	MGMT	Index
NL1	1.268	1.254	0.575	0.947	1.379	0.600	0.737	0.752	1.652	0
NL2	1.528	1.271	1.406	1.406	1.278	1.268	1.803	0.869	0.322	0
NL3	1.127	1.823	1.008	0.860	0.630	1.638	1.932	1.567	1.408	0
NL4	0.347	0.561	0.836	0.556	1.113	0.629	0.702	0.496	0.982	0
NL5	1.321	0.614	1.466	1.571	0.809	1.276	0.555	1.970	1.360	0
HV219	2.570	1.724	2.813	2.518	2.180	1.134	0.966	0.752	0.798	0
HV229	1.081	0.610	0.508	1.030	2.402	0.928	2.378	1.020	1.448	0
HV238	0.819	2.542	0.836	1.684	3.956	0.694	1.035	1.238	0.729	1
HV239	0.848	1.011	0.154	2.045	0.989	0.092	1.149	1.229	0.781	2
LC224	0.494	0.868	0.369	1.805	2.241	0.638	2.313	0.482	1.136	0
LC232	4.323	1.900	1.241	1.058	3.444	1.368	2.928	1.622	2.539	2(1)
LC241	0.372	1.054	0.247	0.490	2.721	0.544	2.549	0.822	1.287	1
LC243	1.793	1.004	0.162	1.696	1.964	0.377	0.563	0.503	0.552	1
LC247	3.487	0.950	0.421	2.793	2.369	0.656	0.514	0.701	1.024	1
LC248	6.507	2.900	0.928	5.322	5.148	0.961	1.338	1.715	1.607	3(1)
LC250	4.234	4.744	1.285	3.463	2.180	1.008	1.444	0.846	0.580	3(2)
LC253	1.507	0.833	0.656	3.299	2.210	0.342	3.249	0.668	1.010	2(1)
LC254	3.309	0.419	0.037	1.349	1.541	0.114	0.620	0.332	0.347	3
LC255	0.194	0.394	0.027	0.501	1.038	0.067	0.532	0.133	0.416	4
LC256	0.708	0.337	0.259	0.374	7.180	0.080	1.347	1.255	0.639	3(2)
LC259	7.579	8.433	1.073	2.132	5.329	2.238	1.625	0.945	0.942	3(2)
120N2	0.427	0.176	0.181	0.477	1.235	0.385	1.828	0.379	0.576	2
121N2	0.903	1.554	0.616	0.552	2.683	0.286	1.765	0.846	0.416	1
122N2	0.467	0.192	0.629	1.096	3.795	0.412	1.932	0.817	1.003	2(1)
123N2	1.921	1.401	1.250	5.177	2.091	0.548	2.445	1.355	0.709	1(0)
124N2	1.989	2.920	0.934	1.030	1.858	1.111	1.404	1.171	1.120	0
125N2	2.553	0.378	0.754	0.679	1.833	0.430	2.969	1.412	0.885	0
127N2	18.791	1.362	0.183	3.842	2.957	0.091	1.292	0.668	0.787	4(3)
128N2	0.418	0.336	0.162	0.341	0.815	0.401	0.559	0.184	0.560	2
130N2	0.354	0.162	0.490	0.764	1.795	0.247	0.973	0.376	0.501	2
131N2	1.073	0.700	0.182	0.537	0.969	0.401	1.670	0.179	1.090	2
132N2	1.233	0.343	0.245	0.567	1.024	0.334	0.946	0.400	0.666	1
133N2	52.783	5.525	1.397	3.737	4.299	0.334	4.438	1.876	1.053	5(3)
134N2	2.813	12.782	0.169	0.723	0.776	0.016	0.390	0.513	0.185	4
137N2	0.442	0.394	0.415	2.161	1.618	0.094	0.753	0.506	0.455	1
143N2	11.096	2.213	1.073	9.660	2.610	0.097	2.099	1.703	1.541	3(2)
145N2	0.385	0.064	0.110	0.575	1.607	0.342	0.629	0.479	0.367	2
146N2	0.093	0.027	0.079	0.374	0.402	0.069	0.257	0.104	0.194	7
120T	190.283	1.501	0.054	0.039	0.017	0.039	0.655	0.358	0.036	6
121T	107.040	5.155	0.396	0.070	0.331	0.209	0.310	0.181	0.091	7
122T	220.098	10.310	2.698	0.304	1.243	1.368	1.866	2.766	0.613	3
123T	3.345	2.578	1.312	0.004	0.898	0.595	1.986	0.999	1.067	2
124T	306.980	4.488	0.326	0.006	0.171	0.567	2.848	0.266	0.867	6
125T	0.042	1.491	0.315	0.034	0.074	0.019	0.104	0.432	0.021	6
127T	14.143	2.167	0.211	1.731	1.314	0.012	0.702	0.479	0.322	4
128T	127.292	5.263	2.161	0.263	0.512	0.387	0.503	0.602	0.330	4
130T	234.265	8.550	0.213	0.007	0.073	0.296	0.463	0.204	0.588	7
131T	35.555	6.004	0.208	0.467	0.300	0.382	1.189	0.101	0.357	5
132T	333.606	8.374	0.526	0.903	0.657	0.487	1.558	0.513	0.323	3
133T	576.829	8.791	0.229	0.054	0.326	0.012	0.551	1.535	0.155	7
134T	1.571	18.076	0.161	0.146	0.097	0.034	0.111	0.056	0.075	8
137T	71.606	12.519	0.477	0.003	0.771	1.975	2.114	0.325	0.690	4
143T	3.736	0.672	2.932	4.830	1.053	0.010	0.540	0.726	0.197	4
145T	326.740	2.840	0.216	0.219	0.149	0.522	0.959	0.141	0.320	6
146T	29.081	1.554	0.088	0.003	0.186	0.387	0.277	0.088	0.176	7

3倍量上调用粗体表示, 3倍量下调用斜体表示.

表3 9个基因在各组中的相对表达 (mean±SD)

组织类型	n	GPC3	E2F1	MMP2	CLDN10	HGF	PTEN	DLC1	PRDM2	MGMT
正常	5	1.12±0.45a	1.10±0.52a	1.06±0.38	1.07±0.41	1.04±0.32c	1.08±0.45ae	1.15±0.66	1.13±0.61	1.14±0.52ae
肝炎	4	1.33±0.84	1.47±0.85a	1.08±1.19	1.81±0.63	2.38±1.22a	0.71±0.45	1.38±0.67	1.06±0.23	0.94±0.34a
肝硬化	12	2.87±2.46a	1.99±2.40a	0.56±0.46	2.02±1.48a	3.11±1.83a	0.70±0.63	1.59±0.97	0.83±0.49	1.00±0.62ae
癌旁(2 cm)	12	5.33±12.99a	1.67±3.18a	0.50±0.40	1.53±1.55a	1.82±1.08ace	0.34±0.25	1.49±1.06	0.69±0.50	0.63±0.32
癌	17	151.84±163.51	5.90±4.71	0.74±0.94	0.54±1.19	0.48±1.43c	0.43±0.52	0.98±0.81	0.57±0.68	0.37±0.30
P值		0.000	0.003	0.359	0.045	0.000	0.024	0.407	0.244	0.000

^aP<0.05 vs 癌组织;

^cP<0.05 vs 肝硬化组织;

^eP<0.05 vs 癌旁组.