人成骨样细胞机械牵拉力敏感基因cDNA扣除文库的构建.pdfVIP

# 270 # ( J Pract Stomatol) 2004 May , 20( 3) cDNA = : cDN A : Saos- 2 , 12% , 12 / min , 12 h mRNA , cDNA, P CR , cDN A , : cDNA , 200 , : P CR cDNA = SubstractivecDNA library construction of genes sensitive to mechanical stretch in human osteoblast like cells Feng Xue, Li Yongming, Ouyang Weiming, et al. Department of Orthodontics, College of Stomatology, TheFourth Military Medical niversity, Xi. an 7 10032, China = Abstract Objective: To reconstruct a sub stractive comp lement ary deoxyrib ornucleic acid ( cDNA ) libr ar y of gen es sensitive to mech anical stretch in human ost eoblast like cells. Methods: Mech an ical str etch at 12 cycles p er minut e w as applied to human ost eob last lik e cells Saos- 2 an d t he deformation of t he str etch ed cells was 12% . Tw elv e h ours aft er loading , mRNAs w ere isolat ed from b oth str etched an d un stretched cells. Substrac2 tiv e cDNA libr ar y of th e gen es sensitive to str etch w as construct ed wit h th e t echn ique based on polymerase ch ain r eaction ( PCR) an d substract ive hybr idizat ion. Pr imar y sequ en cing of clon es in t he library was car ried out . Results: A substract ive cDNA library of gen es sensitive to stretch was con stru ct ed with a cap acity of ab out 200 clones. A ccording to the r esu lt s of sequen cing, most genes in th e library were relat ed to th e mech anical stimulation . One n ovel gene fragment was obt ain ed. Conclusion: The met hod used in th e exp er iment is effec2 tiv e in clon ing genes sensitive to mech anical stretch. =Key ords Substractive hybridization; Polymerase chain reaction; Gene library; Os2 teoblast; Mechanical stretch : R7 83. 5 : A : 100123733( 2004)20320270203 , ,