The Detection of Hamster Connexins: A Comparison of Expression Profiles with Wild-Type Mouse and the Cancer-Prone Min Mouse

Figures & data

TABLE 1 Overview of Rattus norvegicus connexins detected and sequenced, and their accession numbers in GenBank

			Lacking to complete ORF^a
Cx	Acc. no.	Length	5′	3′
26	AY233214	531	60	90
29	AY233215^b	240	141	293
39	AF536558^b	634	134	318
45	AF536559	1150	—	42
47	AY233216^b	1228	64	21
50	AF536560^b	300	498	525
57	AY233217^b	1273	54	191

^aNumber of nucleotides according to the corresponding mouse sequence.

^bSequences were also published by other groups during the course of this work.

TABLE 2 Overview of Mesocricetus auratus connexins and accession numbers in GenBank

			Lacking to complete ORF^a		Included of UTRs
Cx	Acc. no.	Length	5′	3′	5′	3′
26	AY570779	628	—	267	214	—
29	AY570780	371	245	152	—	—
30	AY570781	1467	—	—	333	348
30.2	AY570782	601	21	215	—	—
30.3	AY570783	1022	—	179	400	—
31	AY570784	1563	—	—	104	646
31.1	AY570785	908	45	—	—	137
32	AY570786	556	4	292	—	—
37	AY570787^b	1511	3	—	—	512
39	AY570788	703	114	269	—	—
40	AY570789^b	1247	88	—	—	258
43	AY206455^b	1488	—	—	193	146
45	AY570790^b	1538	—	—	240	107
46	AY570791	1216	102	—	—	64
47	AY571376	1136	101	86	—	—
50	AY570792	1302	52	—	—	37
57	AY570793	897	102	516	—	—

^aNumber of nucleotides according to the corresponding mouse sequence.

^bPartial sequences were also published by other groups during the course of this work.

TABLE 3 Overview of Cricetulus griseus connexins and their accession numbers in GenBank

			Lacking to complete ORF^a		Included of UTRs
Cx	Acc. no.	Length	5′	3′	5′	3′
26	AY286255	531	60	90	—	—
30	AY286256	845	—	—	16	43
30.2	AY514675	304	107	426	—	—
30.3	AY514676	1065	—	23	290	—
31	AY514677	1022	3	—	—	218
31.1	AY571377	824	69	—	—	77
32	AY286257	992	33	—	—	173
37	AY514678	1723	—	—	199	522
39	AY514679	640	171	275	—	—
40	AY286258	938	86	53	—	—
43	AY206456^b	1477	—	—	190	138
45	AY514680	1296	—	—	1	104
46	AY571378	1127	102	25	—	—
47	AY286259	1270	26	27	—	—
50	AY514681	1169	52	96	—	—
57	AY514682	1235	39	244	—	—

^aNumber of nucleotides according to the corresponding mouse sequence.

^bPartial sequence was also published by another group during the course of this work.

TABLE 4 Primer-combinations for screening of connexin expression in Syrian hamster (Mesocricetus auratus) organs and cells, V79 cells (Chinese hamster, Cricetulus griseus) and mouse (Mus musculus, Mm) organs

Cx	Primer combination^a	Primer sequences^b	Size	Conditions (n × X)^c
Ha26	huCx26-40U21 mCx26-437L23	AACAARCACTCCACCAGCATT AAGAYGACCCGGAAGAAGATGCT	398	37 × 65°C
Mm26	Cx26-449U21 Cx26-716L20	TTAAGGACATCGAAGAGATCA AGAATGCAAATTCCAGACAC	268	32 × 56°C
29	mCx29-224U23 mCx29-648L22	CGYTTCTGGGYCTTCCARGTCAT AAACCCARAAGCRCAAGCTCCA	425	35 × 63°C
Ha30	sm30-495U23 30-3UTR-3L22h	GGTACTGAAATGTGGCATTGAGC ACTCTAGACCTAGAGGGAGTTG	384	35 × 62°C
Mm30	h_b6_Cx30_466U20 h_b6_Cx30_617L20	TTCCTTTACAATGGGTACCA TTAAGCAGCATGCAAATCAC	152	32 × 56°C
30.2	m_Cx30.2_2U20 h/m302-168L21	TGGGGGAGTGGGCGTTCCTA CGTGTTRCACACGAACTCCTC	169	8 × 70–63°C + 29 × 62°C^d
30.3	mCx303-114U24 mCx303-743L22	AGCCARCATGAACTGGGSATTTCT TCCATGCAYCTCTTGCCCACCA	651	35 × 65°C
Ha31	sh/mCx31-719U22 31-3UTR-1L23sh	TCTCACCATCTGTGAGATCTKC ACCAGCTTAGCTAAGCAGACCCT	440	35 × 64°C
Mm31	Cx31-182U20 Cx31-621L20	GCGCATCTGGCTGTCAGTAG CTATGCTGGCGCACTGTACC	440	30 × 64°C
Ha31.1	s/m311-550U25 v/hu311-1211L23	CTCTACGTATTCCATGTGTTCTACC GGAAAYBGAGGTDCAGAABAGAG	590	35 × 60°C
Mm31.1	huCx311-158U21 rCx311-1303L24	TGRGTCCACCATGAACTGGAG YTCTCACCCTGGGYCACACAGGAA	1134	5 × 65–63°C + 30 × 63°C^d
Ha32	huCx32-45U22 ha/huCx32-441L24	AGGTGTGAATGAGGCAGGA TGATGTGCACCTTGTGCCTCTTCACCT	406	35 × 62°C
Mm32	hCx32-357U27 hCx32-644L22	CAACACATAGAGAAGAAAATGCTACGG CATGAAGACGGTGAAGACGGTT	288	32 × 62°C
Mm33	rCx33-16U21 rCx33-707L24	GCCTTACACCAGCTCCTAGAA GCCTTAAATAAGACATAGAACAGC	689	35 × 60°C
Mm36	m36-118U20 r36-870L22	ATTGTRGGGGAGACGGTGTA TAGAYTGACTTYCTCTTGGCCT	770	35 × 65°C
37	rnCx37-50U24 mCx37-520L24	ACTCGACCGTGGTGGGYAAGATCT ACTGGCCATAGAGGAAGCCTGCCT	471	35 × 70°C
Ha39	h/m39-432U24 v/m39-848L20	CTTCTCTGTGCCCAACCGAGTTTC CATCTTGGTCTGAATGYTCG	420	35 × 62°C
Mm39	m_Cx39_115U20 m_Cx39_299L20	TTGGCAGGGTCACCTATCTA CCGCGGTGTAGGGTGTAAGT	185	32 x 60°C
Ha40	huCx40-66U21 ha/huCx40-741L23	CAAGGTCTGGCTCACYGTCCT CTTATCCATACCTTGCCGGGACT	670	35 × 65°C
Mm40	Cx40-444U18 Cx40-821L21	CAGGGCACCCTACTCAAC TVTTTAGGCACTGATTGAAGT	382	30 × 58°C
Ha43	cha/huCx43-616U26 huCx43-1120L21	TACGGGATTGAAGAGCACGGCAAGGT TTGCTTGTTGTAATTGCGGCA	504	35 × 65°C
Mm43	RnCx43-350U20 RnCx43-730L19	CTGCCTTTCGCTGTAACACT CGCTCAAGCTGAACCCATA	399	35 × 58°C
Ha45	huCx45-28U21 Cx45_604L20	CTAGAGGAGATYCACAACCAT ACATAAAACGGGTGGACTTG	596	35 × 60°C
Mm45	Cx45-468U20 L: As for Ha45	CCAACCMAAACCTAAGCATG	156	30 × 58°C
Ha46	v/m46-848U23 s/m46-1457L23	CCTACGTCTTCAACATCATCTTC TAGATGGCCAAGTCACCHGGTCT	793	35 × 64°C
Mm46	hu-Cx46-585U24 mo-Cx46-1102L24	CAGGCCCACRGAGAAGACCATCTT CCTGCTTGAGCTTCTTCCAGCCCA	113	34 × 67°C
Ha47	mCx47-234U22 moCx47-861L22	TGCGCTTCTGGGTCTTCCARAT GCCATCTCACAGAGGTTGAGCA	631	35 × 65°C
Mm47	mCx47-56U22 L: As for Ha47	AAYCAYTCCACCTTCGTGGGCA	806	37 × 65°C
50	h_Cx50_455U20 h_Cx50_794L20	GCCACATCATCTTCAAGACC TAGCCCTTGGCTTTCTGGAT	340	35 × 60°C
Ha57	huCx62-17U23 hCx62-781L22	TAYTGGGTGGCATCCTAGAGGAA AATGGAAYGGRGGGCCTGTTTC	765	35 × 66°C
Mm57	moCx57-894U26 huCx57-1034L22	GATAGGCCAATATATTCTCTATGGGT GCAATGCTGTGCATRAAAAGCA	141	11 × 70–60°C+ 25 × 60°C^d

^aThe upper line of primer set indicates the upstream (U) primer, and the lower line indicates the downstream (L) primer. Otherwise, the naming of the primers does not necessarily follow the numbering found in any entry of any database.

^bDegenerations: R = A + G; Y = C + T; S = C + G; K = G + T; M = A + C; B = C + G + T; D = A + G + T; H = A + C + T; V = A + C + G.

^cStandard PCR conditions: 95°C for 5 min, (95°C for 45 sec, X°C for 30 sec, 72°C for 1 min) for n cycles, then 72°C for 7 min.

^dTouch down. For Cx30.2 the method should be read as: After first heating at 95°C for 5 min, the samples were run through eight cycles with annealing temperature decreasing 1°C per cycle from 70°C to 63°C. This was followed by 29 standard cycles at 62°C. The other touch-down methods are read in the same manner.

In all cases, the same primers were used for the two hamster species, and are designated by Ha in the Cx column. Where species designation is not used, common primers were used for hamsters and mice.

TABLE 5 Expression of connexins in some Syrian hamster organs, SHE cells and V79 cells as detected by RT-PCR

Cx	Brain	Eye	Heart	Kidney	Liver	Testis	SHE	V79
26	++	++		++	++	++
29	++		?			++	?
30	++	++		?		++
30.2	++						++	++
30.3		++		++		++	++	++
31	++	++				++	++	++
31.1				?			++	++
32	++	++		++	++
37	++	++	++	++	++	++		++
39		?		?		++
40	++	++	++	++	++	++		++
43	++	++	++	++		++	++	++
45	++	++	++	++	++	++	++
46		++
47	?			++
50		++				++		++
57					++

++: Strong and reproducible expression.

+: Low and reproducible expression.

?: Uncertain (low, but not reproducible).

−: Not expressed.

Figure 1 Similarities and differences in connexin expression in hamster and mouse. Expression of (A) Cx29, (B) Cx39, (C) Cx46, and (D) Cx57 in some hamster organs and cells (left panels) and in some mouse organs (right panels). RNA was isolated from several Syrian hamster and mouse organs, Syrian hamster embryo (SHE) cells, and Chinese hamster V79 cells. The RNA was DNase treated. After removal of DNase, the samples were divided in two. One part was reverse transcribed, the other part was handled similarly but without reverse transcriptase. PCR was then performed on both samples. The upper panels show samples that were reverse transcribed, and the lower panels show the corresponding samples that were not reverse transcribed, indicating the lack of genomic contamination. The positive controls (pos.) are samples that have not been cleaned from genomic contamination. The positions of size markers are indicated to the left.

TABLE 6 Connexin expression profiles in wild-type and Min mouse as detected by RT-PCR

	Wild type					Min
Cx	Brain	Eye	Kidney	Liver	Testis	Brain	Eye	Kidney	Liver	Testis
26	++	++	++	++	++	++	++	++	++	++
29	++	++		++	^a	++	++	?	++
30	++	++	++		++	++	++	++		++
30.2	++		++		++	++		++		++
30.3	?	++	++		^a		++	++
31	?	++	++		++		++	++		++
31.1		++^a	++				++	++
32	++	^a	++	++		++		++	++
33	++	?			++	++	?			++
36	++	++				++	++
37	++	++	++	++	++	++	++	++	++	++
39					++					++
40			++	++	++			++	++	++
43	++	++	++	++	++	++	++	++	++	++
45	++	++	++		++	++	++	++		++
46	++^a	++	++^a		++^a	?	++	+++		++
47	++	?	^a			++	?
50	?	++			++	?	++			++
57		++		++	++		++		++	++

++: Strong and reproducible expression.

+: Low expression.

?: Uncertain (low, but not reproducible).

−: Not expressed.

^a: Evidently different expression level compared with hamster.