Experimental Research of Circular RNA hsa_circ_0000254 in Acute Myeloid Leukemia / 中国实验血液学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To clone the circular RNA hsa_circ_0000254 and construct its lentiviral over-expression vector.</p><p><b>METHODS</b>The sequence of hsa_circ_0000254 (a total of 524 bp long) was synthesized and cloned by using pGH vector. The vector was cut by EcoR I and BamH I, and artificial hsa_circ_0000254 was obtained, then inserted in pLCDH-ciR to construct the recombinant expression vector pLCDH-circ254(C254), which was confirmed by DNA sequencing. The lentiviral expression vectors pLCDH-circ254(C254) and NC(pLCDH-ciR) were cotransfected into 293T cells by lipofectamine 2000(lipo2000). After transfection for 40 hours, the cells were collected and verified by PCR and sequencing.</p><p><b>RESULTS</b>Restriction analysis and DNA sequencing demonstrated that the lentiviral vector pLCDH-circ254(C254) was constructed successfully, the expression efficiency increased 10000 times after transfection of cells.</p><p><b>CONCLUSION</b>The successful construction of the lentiviral expression vector pLCDH-circ254(C254) results in the production of high-titer lentivirus, so as to facilitate further study of the molecular functions of hsa_circ_0000254.</p>