1 / 11

GFP silencing reverted cells

Figure 1A. Vectors employed. Control line. Experimental line-1. Experimental line-2. GFP silencing reverted cells. Sf21 cell. GFP expressing cells. X. GFP silenced cells. Figure 1B. gfp shRNA Forward primer. gfp shRNA (49 bp). a. EcoRI. KpnI. 19 bp. 9 bp. 19 bp. gfp shRNA

eileen
Download Presentation

GFP silencing reverted cells

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Figure 1A Vectors employed Control line Experimental line-1 Experimental line-2 GFP silencing reverted cells Sf21 cell GFP expressing cells X GFP silenced cells

  2. Figure 1B gfp shRNA Forward primer gfp shRNA (49 bp) a. EcoRI KpnI 19 bp 9 bp 19 bp gfp shRNA Reverse primer 1 2 3 e. i 1 2 3 4 b. 6000 PCR d. f. 1 M 2 1 2 3 4000 1 2 23 nt 2000 c. i RT-PCR 21 nt 0 -actin loading control ii mRNA ii Total RNA loading control 14 kDa iii

  3. Visible Light UV Light Figure 2 A i B 200 160 ii 120 Counts 80 iii 40 0 101 100 102 103 104 FLH-1

  4. Figure 3 FHVB2 expressing line GFP expressing line GFP silencing line 2400 FHVB2- C17 line (i) 1600 800 Relative intensity of siRNA 0 1 2 3 4 (ii) siRNA (iii) Total RNA loading control

  5. Figure 4 A 1 2 3 M kDa C B 15 13 14 15 16 17 18 mAU Catalase 16 14 (i) 1600 1400 160 (ii) 1200 1000 (iii) 110 800 Aldolase 73 600 400 200 E M 1 2 0 0.0 5.0 10.0 15.0 20.0 ml 23mer D 10 11 12 13 14 15 16 17 18 19 20 21 22 23 21mer

  6. Figure 5 A 15 16 Pool W. Probe B 15 i 16 - FHVB2 14 mAU 1600 ii -DCR-1 PAZ 1400 C 1200 10 11 12 13 14 15 16 17 1000 800 18 19 20 21 22 23 +ve 600 400 200 0 0.0 5.0 10.0 15.0 20.0 25.0 ml

  7. Figure 6 A (i) (ii) (iii) (iv) (vi) (v) AD(PAZ) BD(FHV B2) AD(ENDO) BD(FHV B2) pGBKT7 FHVB2 pGADT7+ pGBKT7 pGBKT7 AH109 AD (REP) +BD (REP) pGADT7 SD (Leu-Trp-His-) Scheme YPD SD (Trp-) SD (Leu-) SD (Leu- Trp-) B (ii) (i) INPUT ELUTES FLOW THROUGH I (iii) FT EL M 1 2 3 M 1 2 3 1 2 3 4 M 5 6 116 66 45 35 25 18

  8. Figure 7 A pGAD + pGBD AH109 pGBD (FHVB2) pGAD AD(DCR2PAZ) +BD (FHVB2) AD(Rep)+ BD(Rep) AD (DCR2 PAZ)+ pGBD pGBD (i) Scheme (ii) SD (Leu-Trp-) (iii) SD (Leu-Trp-His-) GST-FHVB2 B MBP MBP-DCR-2-PAZ INPUT FT EL EL FT kDa M 1 2 3 4 5 6 7 118 81 52 36

  9. Figure 8 N C A Full Length FHVB2 (107aa) N C FHVB2 – C17 (90aa) B ( ) % ( ) ( ) % % Sf21 cell line GFP expressing GFP silenced C 1.2 1.0 Relative level of gfp mRNA 0.8 0.6 ( ) % ( ) % 0.4 0.2 0 GFP GFP silenced GFP reverted FHVB2- C17 FHVB2- GFP reverted FHVB2- C17 transfected

  10. Figure 9 A AH109 AH109 AD+BD AD+BD AD AD BD(FHV-B2) BD(FHV-B2) AD(PAZ) DCR1+BD (FHVB2) AD(PAZ) DCR1+BD (FHVB2) AD(REP) +BD(REP) AD(REP) +BD(REP) AD- DCR1 (PAZ)+ BD(FHVB2-C17) AD- DCR2 (PAZ)+ BD(FHVB2-C17) BD BD (ii) SD (Leu-Trp-His-) (iii) Scheme (iv) SD (Leu-Trp-His-) (i) Scheme B (i) (ii) INPUT F T ELUTE FT INPUT ELUTE 4 5 M 6 7 1 2 M 3 kDa M 1 2 3 118 81 52 36

  11. Supplementary Figure 1 dm Dicer-1-PAZ dm Dicer-2-PAZ A. dm Dicer-1 dm Dicer-1-PAZ dm Dicer-2-PAZ dm Dicer-1-PAZ dm Dicer-2-PAZ B. 1715 amino acids N-terminal C-terminal dm Dicer-2 dsRNA bd RNase III dsRNA bm HELICASES PAZ MBP-ENDONUCLEASE MBP-PAZ 1710aa 1aa 554aa 659aa 838aa 945aa 1187aa 1642aa 1646aa 535aa MBP-PAZ C. 2249 amino acids N-terminal C-terminal dsRNA bd RNase III HELICASE-C DUF 283 PAZ 2241 2175aa 544aa 510aa 606aa 825aa 920aa 1096 aa 1218 aa 1706 aa 1919 aa aa

More Related