Construction of the subtracted cDNA library of striatal neurons treated with long-term morphine / 中国医学科学杂志(英文版)
Chinese Medical Sciences Journal
;
(4): 54-59, 2011.
Article
Dans Anglais
| WPRIM
| ID: wpr-299413
ABSTRACT
<p><b>OBJECTIVE</b>To construct a morphine tolerance model in primarily cultured striatal neurons, and screen the differentially expressed genes in this model using suppression subtractive hybridization (SSH).</p><p><b>METHODS</b>Sbtracted cDNA libraries were constructed using SSH from normal primarily cultured striatal neurons and long-term morphine treated striatal neurons (10-5 mol/L for 72 hours). To check reliability of the cell culture model, RT-PCR was performed to detect the cAMP-responsive element-binding protein (CREB) mRNA expression. The subtracted clones were prescreened by PCR. The clones containing inserted fragments from forward libraries were sequenced and submitted to GenBank for homology analysis. And the expression levels of genes of interest were confirmed by RT-PCR. Results CREB mRNA expression showed a significant increase in morphine treated striatal neurons (62.85 ± 1.98) compared with normal striatal neurons (28.43 ± 1.46, P < 0.01). Thirty-six clones containing inserted fragments were randomly chosen for sequence analysis. And the 36 clones showed homology with 19 known genes and 2 novel genes. The expression of 2 novel genes, mitochondrial carrier homolog 1 (Mtch1; 96.81 ± 2.04 vs. 44.20 ± 1.31, P < 0.01) and thymoma viral proto-oncogene 1 (Akt1; 122.10 ± 2.17 vs. 50.11 ± 2.01, P < 0.01), showed a significant increase in morphine-treated striatal neurons compared with normal striatal neurons.</p><p><b>CONCLUSIONS</b>A reliable differential cDNA library of striatal neurons treated with long-term morphine is constructed. Mtch1 and Akt1 might be the candidate genes for the development of morphine tolerance.</p>
Texte intégral:
Disponible
Indice:
WPRIM (Pacifique occidental)
Sujet Principal:
Pharmacologie
/
Physiologie
/
Données de séquences moléculaires
/
Banque de gènes
/
Cellules cultivées
/
Rat Wistar
/
Corps strié
/
RT-PCR
/
Analyse de profil d'expression de gènes
/
Biologie cellulaire
Limites du sujet:
Animaux
langue:
Anglais
Texte intégral:
Chinese Medical Sciences Journal
Année:
2011
Type:
Article
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