shrna psicor ef1a mch puro vector  (Addgene inc)

Journal: Investigative Ophthalmology & Visual Science

Article Title: Shootin1 Regulates Retinal Ganglion Cell Neurite Development: Insights From an RGC Direct Somatic Cell Reprogramming Model

doi: 10.1167/iovs.65.6.41

Figure Lengend Snippet: Shootin1 is specifically expressed in reprogrammed iRGCs in vitro. Confocal images of iRGCs stained for shootin1 ( red ), TUJ1 ( green , indicating successfully reprogrammed iRGCs), and nuclei (DAPI, blue ). The area outlined by the dashed line in A is shown at a higher magnification in B , and the growth cone area outlined by the dashed line in B is shown at a higher magnification in C . The white arrows in A point to MEFs that were not infected with ABI lentivirus. The yellow arrowheads in A point to TUJ1 + successfully reprogrammed iRGCs. The arrows in C point to the tip of the filipodia where shootin1 accumulated. Scale bars: 100 µm ( A ), 50 µm ( B ), and 5 µm ( C ).

Article Snippet: Short-hairpin RNAs (shRNAs) targeting Shtn1 were cloned into the pSicoR lentivirus vector (Addgene, Watertown, MA, USA).

Techniques: In Vitro, Staining, Infection

Journal: Investigative Ophthalmology & Visual Science

Article Title: Shootin1 Regulates Retinal Ganglion Cell Neurite Development: Insights From an RGC Direct Somatic Cell Reprogramming Model

doi: 10.1167/iovs.65.6.41

Figure Lengend Snippet: Shtn1 deficiency retards the growth of iRGC axons. ( A ) IF images showing that shRNAs targeting Shtn1 abolished the expression of shootin1 in iRGCs. The arrows point to iRGCs that were not infected with shRNA-expressing lentivirus (indicated by lack of GFP expression), and the arrowheads point to iRGCs expressing shRNAs (indicated by GFP expression). ( B ) IF images of TUJ1-stained iRGCs from the control and Shtn1 -knockdown groups at DAI 5. ( C ) A plot showing the measurement results of the length of iRGC axons in B . Numbers inside the brackets indicate the number of iRGCs measured. ( D ) IF images of TUJ1-stained iRGCs from the control and Shtn1 -knockdown groups at DAI 13. ( E ) A plot showing the measurement results of the length of iRGC axons in D . Numbers inside the brackets indicate the number of iRGCs measured. ** P < 0.01 (one-way ANOVA was used to compare each shRNA group with the control group). Scale bars : 50 µm ( A ) and 100 µm ( B , D ).

Article Snippet: Short-hairpin RNAs (shRNAs) targeting Shtn1 were cloned into the pSicoR lentivirus vector (Addgene, Watertown, MA, USA).

Techniques: Expressing, Infection, shRNA, Staining

Journal: Investigative Ophthalmology & Visual Science

Article Title: Shootin1 Regulates Retinal Ganglion Cell Neurite Development: Insights From an RGC Direct Somatic Cell Reprogramming Model

doi: 10.1167/iovs.65.6.41

Figure Lengend Snippet: Shtn1 is required for the iRGC reprogramming process. ( A ) Upper panels : IF images of MEFs stained for ASCLI ( red ) and GFP ( green , indicating control empty vector or shRNA-expressing lentivirus infection) at DAI 2 to show the virus infection efficiency. Middle panels : IF images of reprogramming cells stained for TUJ1 ( red ) and GFP ( green ) at DAI 5. Lower panels : IF images of reprogramming cells stained for TUJ1 ( red ) and GFP ( green ) at DAI 13. Scale bar : 100 µm. ( B ) A plot showing the TUJ1 + iRGC induction efficiency in the control and Shtn1 -knockdown groups at DAI 5. ( C ) A plot showing the TUJ1 + iRGC induction efficiency in the control and Shtn1 -knockdown groups at DAI 13. The data are presented as means ± SD, corresponding to three independent biological replicates. ** P < 0.01 (three biological replicates, Student's t -test).

Article Snippet: Short-hairpin RNAs (shRNAs) targeting Shtn1 were cloned into the pSicoR lentivirus vector (Addgene, Watertown, MA, USA).

Techniques: Staining, Plasmid Preparation, shRNA, Expressing, Infection, Virus