Generation of stable transgenic embryonic stem (ES) cell lines by classic transfection is still a difficult task, requiring time-consuming clonal selection, and hampered by clonal artifacts and gene silencing. Here we describe a novel system that allows construction of lentivectors and generation of stable ES cell lines with > 99% transgene expression within a very short time frame. Rapid insertion of promoters and genes of interest is obtained through a modular recombinational cloning system. Vectors contain central polypurine tract from HIV-1 element and woodchuck hepatitis virus post-transcriptional regulatory element as well as antibiotic resistance to achieve optimal and homogenous transgene expression. We show that the system 1) is functional in mouse and human ES cells, 2) allows the generation of ES cells expressing genes of interest under the control of ubiquitous or tissue-specific promoters, and 3) allows ES cells expressing two constructs through selection with different antibiotics to be obtained. The technology described herein should become a useful tool in stem cell research.
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Barberi, T
;
Klivenyi, P
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Klivenyi, P
;
Calingasan, NY
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Calingasan, NY
;
Lee, H
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Lee, H
;
Kawamata, H
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Kawamata, H
;
Loonam, K
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Loonam, K
;
Perrier, AL
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Perrier, AL
;
Bruses, J
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Bruses, J
;
Rubio, ME
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Rubio, ME
;
Topf, N
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Topf, N
;
Tabar, V
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Tabar, V
;
Harrison, NL
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Harrison, NL
;
Beal, MF
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Beal, MF
;
Moore, MAS
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Moore, MAS
;
Studer, L
论文数: 0引用数: 0
h-index: 0
机构:
Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USACornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Barberi, T
;
Klivenyi, P
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Klivenyi, P
;
Calingasan, NY
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Calingasan, NY
;
Lee, H
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Lee, H
;
Kawamata, H
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Kawamata, H
;
Loonam, K
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Loonam, K
;
Perrier, AL
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Perrier, AL
;
Bruses, J
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Bruses, J
;
Rubio, ME
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Rubio, ME
;
Topf, N
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Topf, N
;
Tabar, V
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Tabar, V
;
Harrison, NL
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Harrison, NL
;
Beal, MF
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Beal, MF
;
Moore, MAS
论文数: 0引用数: 0
h-index: 0
机构:Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA
Moore, MAS
;
Studer, L
论文数: 0引用数: 0
h-index: 0
机构:
Cornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USACornell Univ, Weill Med Coll, Div Neurosurg, Lab Stem Cell & Tumor Biol, New York, NY 10021 USA