scholarly journals Using recombinase‐mediated cassette exchange to engineer MIN6 insulin‐secreting cells based on a newly identified safe harbor locus

2021 ◽  
Author(s):  
Asami Furukawa ◽  
Aya Tanaka ◽  
Suguru Yamaguchi ◽  
Minami Kosuda ◽  
Midori Yamana ◽  
...  
Keyword(s):  
Safe Harbor ◽  
Cassette Exchange ◽  
Insulin Secreting Cells

scholarly journals Identification of the safe harbor locus, AAVS1, from porcine genome and site-specific integration of recombinase-mediated cassette exchange system in porcine fibroblasts using CRISPR/Cas9

2021 ◽  
Author(s):  
Choongil Lee ◽  
Soo-Young Yum ◽  
Woojae Choi ◽  
Seokjoong Kim ◽  
Goo Jang ◽  
...  
Keyword(s):  
Cell Line ◽  
Pcr Analysis ◽  
Safe Harbor ◽  
Critical Approach ◽  
Porcine Genome ◽  
Transgenic Pigs ◽  
Site Specific ◽  
Donor Vector ◽  
Cassette Exchange ◽  
Aavs1 Locus

Gene integration at site-specific loci , such as safe harbor regions for s table expression via transgenesis ,  is a critical approach for understanding the function of a gene in cells or animals. The AAVS1 locus is a  well-known safe harbor site  for human and mouse studies. In the present study, we found an AAVS1-like sequence in  the porcine genome using  the UCSC Genome Browser and designed TALEN and CRISPR/Cas9 to target AAVS1. The efficiency of CRISPR/Cas9 for targeting the AAVS1 locus in porcine cells was superior to that of TALEN. An AAVS1-targeting donor vector containing GFP was designed and cloned. We added a loxP-lox2272 cassette sequence to the donor vector for further exchange of various transgenes in the AAVS1-targeted cell line. The donor vector and CRISPR/Cas9 components targeting AAVS1 were transfected into a porcine fibroblast cell line. Targeted cells of CRISPR/Cas9-mediated homologous recombination were identified by antibiotic selection. Gene knock-in at the AAVS1 locus was confirmed by PCR analysis. To induce recombinase-mediated cassette exchange (RMCE), another donor vector containing the loxP-lox2272 cassette and inducible (Tet-on) Cre recombinase was cloned. The Cre-donor vector was transfected into the AAVS1-targeted cell line, and RMCE was induced by adding doxycycline to the culture medium. RMCE in porcine fibroblasts was confirmed using PCR analysis. In conclusion, gene targeting at the AAVS1 locus and RMCE in porcine fibroblasts was successful. This technology will be useful for future porcine transgenesis studies and  the generation of  stable transgenic pigs.

When are Firms Sued for Qualitative Disclosures? Implications of the Safe Harbor for Forward-Looking Statements

2019 ◽  
Vol 95 (1) ◽  
pp. 31-55 ◽  
Author(s):  
Richard A. Cazier ◽  
Kenneth J. Merkley ◽  
John S. Treu
Keyword(s):  
Legal Liability ◽  
Safe Harbor ◽  
Litigation Risk ◽  
Securities Laws ◽  
Forward Looking

ABSTRACTPrior research finds that positive tone in firms' qualitative disclosures increases the risk of shareholder lawsuits. However, federal securities laws provide a safe harbor intended to shield firms' forward-looking statements from legal liability. One implication of this safe harbor is that litigation risk potentially varies between qualitative forward- and non-forward-looking statements. Consistent with this implication, we find that positive tone in forward-looking qualitative statements is significantly less related to the likelihood of subsequent litigation than is positive tone in non-forward-looking qualitative statements. On average, we fail to find a significant association between qualitative forward-looking statements and subsequent litigation. We do find evidence, however, that positive tone in qualitative forward-looking statements relates positively to subsequent litigation in two U.S. circuits in which court rulings reduced safe harbor protections for forward-looking statements. Overall, our results are consistent with the safe harbor effectively shielding firms' qualitative forward-looking statements from litigation risk.

scholarly journals Establishing isogenic inducible cell lines using founder reporter lines and recombinase-mediated cassette exchange

BioTechniques ◽  
2013 ◽  
Vol 55 (5) ◽  
Author(s):  
Bin Guan ◽  
Tae Magomi ◽  
Tian-Li Wang ◽  
Ie-Ming Shih
Keyword(s):  
Cell Lines ◽  
Cassette Exchange

scholarly journals Insulin-secreting cells derived from embryonic stem cells normalize glycemia in streptozotocin-induced diabetic mice

Diabetes ◽  
2000 ◽  
Vol 49 (2) ◽  
pp. 157-162 ◽  
Author(s):  
B. Soria ◽  
E. Roche ◽  
G. Berna ◽  
T. Leon-Quinto ◽  
J. A. Reig ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Embryonic Stem ◽  
Diabetic Mice ◽  
Insulin Secreting Cells

scholarly journals Rhesus iPSC Safe Harbor Gene-Editing Platform for Stable Expression of Transgenes in Differentiated Cells of All Germ Layers

2017 ◽  
Vol 25 (1) ◽  
pp. 44-53 ◽  
Author(s):  
So Gun Hong ◽  
Ravi Chandra Yada ◽  
Kyujoo Choi ◽  
Arnaud Carpentier ◽  
T. Jake Liang ◽  
...  
Keyword(s):  
Gene Editing ◽  
Stable Expression ◽  
Safe Harbor ◽  
Germ Layers ◽  
Differentiated Cells

scholarly journals Utilization of the AAVS1 safe harbor locus for hematopoietic specific transgene expression and gene knockdown in human ES cells

2014 ◽  
Vol 12 (3) ◽  
pp. 630-637 ◽  
Author(s):  
Amita Tiyaboonchai ◽  
Helen Mac ◽  
Razveen Shamsedeen ◽  
Jason A. Mills ◽  
Siddarth Kishore ◽  
...  
Keyword(s):  
Transgene Expression ◽  
Es Cells ◽  
Gene Knockdown ◽  
Safe Harbor ◽  
Human Es Cells
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