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Abstract: 6 0fx0f( f  \5Pk ( C ^y  <#2_ж_ж ?=K  zNumber of copies, germ line transmission, and expression level of a GFP transgene in mice generated by lentiviral vectors({08fA{Hf {15Pk ( C ^y(H !#h%0'(*,P.013p5879:<X> @ACxE@G  <_ж_ж ? 8   Teixeira M.*1, Ngre D.*2,3, Blanquier B.4, Ghittoni R.5, Szecsi J.3 , Henry F.1, Coupet CA.5, Aubert D.1, Cosset FL3, Marvel J.5*  " *  " * " *  " *  " *  " *  " *  " *  " *  " * T0 '5Pk ( C ^y(H !#h%0'(*,P.013p5T  <D_ж_ж ? `4' 1.PBES,Ecole Normale Suprieure de Lyon, 2.Plateau de vectorologie - Ecole Normale Suprieure de Lyon, 3.Unit U758 Inserm  ENS Lyon, 4.Plateau de QPCR  CERVI- Lyon, 5.Unit U851, INSERM Lyon * Both authors contributed equally to this work ( # 1$5Pk ( C ^y(H !#h%0'(*,P.0  <|/_ж_ж ?0`$ ,Abstract: Up to now transgenic mice were mainly obtained by pronuclear microinjection of DNA. However, recently an alternative method based on the use of lentiviral vector has been developped, providing better efficiency of trangenesis. The aim of this work was to genetically characterize the founders and their progeny obtained by this new method. Also, we tried to correlate the expression level obtained using three ubiquitous promoters - CMV early enhancer/chicken  actin hybrid promoter (CAG), spleen focus-forming virus promoter (SFFV) and ubiquitin C promoter (FG12) - in blood circulating lymphocytes, with the number of integrated copies (Fig 4). Lentiviral vectors carrying a GFP transgene were injected into the perivitelline space of fertilized oocytes, resulting in 52% to 80% of founder animals carrying the transgene, according to the titre of lentivirus vector preparations. The transmission of transgene to offspring by founders was also analysed. We used quantitative PCR to determine the number of copies of integrated transgene. The number of copies varies between 0.1 and 50, and is directly correlated with the concentration of the lentiviral vectors (Table 1). We found that the founders harbouring less than one copy are mosaics, indeed, all their transgene positive progeny harboured 1 copy (Fig 2). These results indicate that viral integration can occur at 2 and 4 cells stages. Mosacisme was not restricted to low viral titre as some F1 mice present a higher number of integrated copies than their founder. These results suggest that most founders obtained by this lentiviral transduction method, even those with many integrated copies, could be mosaics. GFP expression stability over generation was analyzed by flow cytometry, in peripheral blood lymphocytes of founders or transgenic animals. Expression of GFP seems to be stable during generations. Nevertheless, we noticed, that depending on the insertion site, in certain animals the expression could fade out with time (Fig 3 and fig 4). In conclusion, lentiviral transduction allows to quickly obtain a large number of transgenic mice, widely different genetically in regard of the transgene integration. However, mosaicisme seems to be a common feature of this approach and should be taken into account when analysing founder mice.  0fx0f(  ff" 0   d5Pk ( C ^y(   </_ж_ж ? %pM3 Materials and Methods 1.Generation of transgenic mice by perivitelline injection of fertilized oocytes. 2.Extraction of DNA 3.Determination of copies number by quantitative PCR 4.Blood samples collection 5.Extraction of lymphocytes 6.Determination of GFP fluorescence in lymphocytes b0f  ' f# #  \  :Up  - H c~ ; Bu8 5&  5&TB   c $D@8?5 5 T   c $$?5&x@ &-   &- @ &*   &* T   c $ ?&>'f2   6F81?&&(f2   6f381?''f2   6ĚH;e81?/'In'kf2   6ĚH;e81?'f'TB  c $D81?*()6TB  c $D81?>(,*Rf2   6f381?g''f2   6H?&( 0   B<_ж_ж ?*-s microinjection Z5Pk ( C ^y0   BS_ж_ж ?B)+^ 25% Tg Z5Pk ( C ^yT   c $ ?%'3&.f2   6̙3f81?%]'u`2 !  081?>&''f2 "  6H?%'!)`B #  0Df?&q&TB $  c $D?&q&f2 %  6F81?]'(u`2 &  081?''(TB '  c $D?(q(TB (  c $D?+(q,(f2 )  6̙3f81?%4'L`2 *  081?"&N&`B +  0Df?S&T&@ &Q(t , &Q(tf2 -  6F81?&R(u`2 .  081?>'u'TB /  c $D?o'p'TB 0  c $D?''ZB 1  s *D$?5'(6'8 2  Bh_ж_ж ?*I+  QPCR n=0.5    Z5Pk ( C ^yl 3  <A ̙3f ?&0Z)l 4  <A $?& Z)l 5  <A $?&Z)Hl 6  <A $?&Z) ZB 7  s *D$?G'H'TB 8  c $D?s&t&0 9  B,V_ж_ж ?(;+ 75% WT Z5Pk ( C ^y1 :  BN_ж_ж ?t't*\ Founder Z5Pk ( C ^yT ;  c $ ?+`2f2 <  6̙3f81? x`2 =  081?+f2 >  6H?O !`B ?  0Df?EsFTB @  c $D?ospf2 A  6̙3f81?N`2 B  081?O`B C  0Df?TB D  c $D?f2 E  6̙3f81?R !)`2 F  081? +Q!`B G  0Df? !c !TB H  c $D?$!c%!f2 I  6̙3f81?| *`2 J  081?j+`B K  0Df?bTB L  c $D?bf2 M  6̙3f81? !*`2 N  081? +!`B O  0Df? b TB P  c $D? b f2 Q  6̙3f81?^ N`2 R  081?O] `B S  0Df?  TB T  c $D?/ 0 ZB U  s *D$?f2 V  6̙3f81?7M`2 W  081?#P`B X  0Df?UVTB Y  c $D?mnZB Z  s *D$?Ps6 [  B _ж_ж ?"# QPCR n=1    Z5Pk ( C ^yl \  <A ̙3f ?]!Jl ]  <A $?]!9/ ^  B$_ж_ж ?$!# 50%WT Z5Pk ( C ^y1 _  B%_ж_ж ?5 B# Founder Z5Pk ( C ^y0 `  B'_ж_ж ?F!># 50% Tg Z5Pk ( C ^yT a  c $ ?L/"/*f2 b  6̙3f81?t/1pF2 c  T@_ж_ж81?/"0 14$0f f Z5Pk ( C ^yf2 d  6H?h/!2f2 e  6̙3f81?12pE2 f  THM_ж_ж81?1"P2 4$0f f Z5Pk ( C ^yf2 g  6̙3f81?.;0`2 h  081?/06 i  BT4_ж_ж ?~1%-3m QPCR n=9    Z5Pk ( C ^yl j  <A ̙3f ?-sh0l k  <A ̙3f ?1s3ZB l  s *D$?0W0|@ "- . m "- . n  @ABCEF ?   @"-7- o  @ABC EF ?     @6-eK- p  @ABCEF ?   @L-a-f q  @ABCEF ?  @a-v- r  @ABCEF ?   @v-- s  @ABCEF ?   @-- t  @ABCEF ?    @-X- u  @ABC EF ?    @--Y v  @ABCEF ?   @- - w  @ABCEF ?   @- - x  @ABC EF ?   @- .H y  @ABCEF ?  @ .H. z  @ABCEF ?  @.4. {  @ABCEF ?  @5.J.r |  @ABCEF ?   @J.q_. }  @ABCEF ?  @_.t.P ~  @ABCEF ?  @t.P.   @ABCEF ?   @..@ ,.  ,.TB   c $D?,.TB   c $D?--TB   c $D?{.|.TB   c $D?F-G-+   BH8_ж_ж ?- 5.  7  Z5Pk ( C ^y,   Bk_ж_ж ?X. R/  14  Z5Pk ( C ^y+   B_ж_ж ?, a-  0  Z5Pk ( C ^y=   BXz_ж_ж ??.{1k   Copy number$ 0f  f  Z5Pk ( C ^y@ 1 3  1 3@ 92 3  92 3   @ABCEF ?   @92N2   @ABC EF ?     @N2fc2   @ABCEF ?   @c2x2f   @ABCEF ?  @x22   @ABCEF ?   @22   @ABCEF ?   @22   @ABCEF ?    @2X2   @ABC EF ?    @22Y   @ABCEF ?   @2 2   @ABCEF ?   @2  3   @ABC EF ?   @ 3!3I   @ABCEF ?  @"3H73   @ABCEF ?  @73L3   @ABCEF ?  @L3a3r   @ABCEF ?   @a3qv3   @ABCEF ?  @v33Q   @ABCEF ?  @3P3   @ABCEF ?   @33@ 13  13TB   c $D?13TB   c $D?22TB   c $D?33TB   c $D?^2_2+   Bx2Z_ж_ж ?2 2  0  Z5Pk ( C ^y+   B0BZ_ж_ж ?2 \3  2  Z5Pk ( C ^y+   BEZ_ж_ж ?b3 4  4  Z5Pk ( C ^yZB   s *D$?1  1Df2   6̙3f81?b0;2`2   081?O1"2r&   <ZZ_ж_ж ?.&0 14 Z5Pk ( C ^y%   <eZ_ж_ж ?1`2 4 Z5Pk ( C ^y1   B(qZ_ж_ж ?&..31 Founder Z5Pk ( C ^y-   B}Z_ж_ж ?Y.$0 50% Z5Pk ( C ^y0   BZ_ж_ж ?!13 50% tg Z5Pk ( C ^yTB   c $D?050F   H Z_ж_ж ?J/./<  newborn number$0f f Z5Pk ( C ^y_   <Z_ж_ж ?u 3& CFig. 1. The oocyte injection is performed at one cell stage. A- If integration occurs at 1 cell stage there is no mosaicism . If single (B) or multiple (C) integrations occur at two cells stage we observe mosaicism. The backcross with wild type mice of founders having 1 GFP integration per genome result in : 50% (A) or 25% (B) of transgenic mice carrying one GFP integration per genome. Backcross to wild type of founders with multiple integrations (C) give rise to some newborns with number of GFP copy per genome that are higher than the one measured for the founder parent. DD DZ5Pk ( C ^yZB   s *D$?N$O$\ZB   s *D$?{,|,/1   BTZ_ж_ж ?t A"0ff Z5Pk ( C ^y1   BPZ_ж_ж ?~$t% B"0ff Z5Pk ( C ^y1   BZ_ж_ж ?-. C"0ff Z5Pk ( C ^yP@ ]+#Z  ]+#ZB  bENFGJSUNVFW$? `T`T0*0*`T`T`$ 0*0*0*0*0*L 0*0*0*0*0*L @0*G 0* @0*G 0* @0*H 0* @0*H 0*/*_T/*!f   6A  ? +#P@ &+  &+B  bENFGJSUNVFW$? `T`T0*0*`T`T`$ 0*0*0*0*0*L 0*0*0*0*0*L @0*G 0* @0*G 0* @0*H 0* @0*H 0*/*_T/*3' ){f   6A  ?6(+P@ E0w4  E0w4B  bENFGJSUNVFW$? `T`T0*0*`T`T`$ 0*0*0*0*0*L 0*0*0*0*0*L @0*G 0* @0*G 0* @0*H 0* @0*H 0*/*_T/*02gf   6A  ?1w4;   B Z_ж_ж ?b(+  Wild Type$ 0   Z5Pk ( C ^y;   BZ_ж_ж ?14  Wild Type$ 0   Z5Pk ( C ^y;   BX  0333?<>>X  0333?<Y??X  0333?D\DX  0333?DD EX  0333?2G0HX  0333?2}HHLB  c $D?<;;=LB  c $D?sBuBLB  c $D?eBBLB  c $D?^FFLB  c $D$? 88LB  c $D$?z88LB  c $D$?3z884LB  c $D$?z88LB  c $D$?<z88=LB  c $D$? z88 L  c $ ?77L  c $ ?@77L  c $ ?77TL  c $ ?77+  B 2 ?7H8 0,5"0 Z5Pk ( C ^yL  c $ ?77L  c $ ?L77L  c $ ?77\L  c $ ?77)  B ? 7.8- 1"0 Z5Pk ( C ^yXB  0D333o?8:XB  0D333o?<<@XB  0D333o?:B~EXB  0D333o?F.IL  c $$?; K6I4+  B$_ж_ж ? z74;  Example 1    Z5Pk ( C ^y+  Bq_ж_ж ? 0>X  0333@8??4@X  0333@8?AALB  c $D@8?88LB  c $D@8?UR:S:VLB  c $D@8???LB  c $D@8?WUAVAXLB  c $D@8?BBLB  c $D@8?&D(DLB  c $D@8?EELB  c $D@8?FFLB  c $D@8?&aHbH'LB  c $D@8?IILB  c $D@8?88LB  c $D@8?UR:S:VLB  c $D@8???LB  c $D@8?BBLB  c $D@8?&D(DLB  c $D@8?EELB  c $D@8?FFLB ! c $D@8?IIX " 0333@8?8N9LB # c $D@8?88LB $ c $D@8?89- % B ?8O9< 8,4$0 Z5Pk ( C ^y. & B\ ?+ :: 11,2$0 Z5Pk ( C ^y. ' B ?d;?<* 10,1$0 Z5Pk ( C ^y. ( BC ?h<= 10,7$0 Z5Pk ( C ^y- ) BLR ?"T>> 8,9$0 Z5Pk ( C ^y. * BE ??n@d 11,9$0 Z5Pk ( C ^y- + B_ ?0AA 9,2$0 Z5Pk ( C ^y8 FoKG , oFKG` -  0333@8?F!5G5 .  Bu ?FoLG 1,4$0 Z5Pk ( C ^y8 GSH / SGH` 0  0333@8?GqH5 1  Bt ?GSH 1,6$0 Z5Pk ( C ^y8 NEE 2 NEE` 3  0333@8?NEE5 4  B ?\EE 2,3$0 Z5Pk ( C ^y8 |B&C 5 |B&C` 6  0333@8?|BC5 7  B ?B'C 9,2$0 Z5Pk ( C ^y8 CcD 8 cCD` 9  0333@8?CqD6 :  BH ?CcD 10,7$0 Z5Pk ( C ^y+ ; Bp ?781 0$0ff Z5Pk ( C ^y+ < Bȳ ?78R 2$0ff Z5Pk ( C ^y+ = B˳ ?78o 4$0ff Z5Pk ( C ^y+ > BHԳ ?78 6$0ff Z5Pk ( C ^y+ ? BH ?78 8$0ff Z5Pk ( C ^y, @ B ?k7 8 10$0ff Z5Pk ( C ^y, A B ?<k7 8 12$0ff Z5Pk ( C ^y, B B@ ?^k7 8  14$0ff Z5Pk ( C ^y< C HL_ж_ж ?67 Copy number / cell  Z5Pk ( C ^y! D Bď_ж_ж ?67 B Z5Pk ( C ^y+ E Bˏ_ж_ж ?=bA  Example 5    Z5Pk ( C ^yL F c $$?'p5p> G <܏_ж_ж ?U95= <jFig. 3: GFP expression in F0, F1 and F2 mice Analysis of GFP expression in blood lymphocytes in three generations (F0, F1, F2) of mice harbouring one GFP copy. Backcross of 25F0 mouse with wild type resulted in 6 transgenic mice with 1 transgene copy: 78F1, 41F1, 36F1, 82F1 and 77F1. All F0 and F1 animals and most of F2 mice express GFP in blood lymphocytes. :k&f=k ^5Pk ( C ^y(L H c $ ?'(p) )LB I c $D$?R384U3LB J c $D$?2842LB K c $D$?0840LB L c $D$?/84/LB M c $D$?j.84k.LB N c $D$?/-842-LB O c $D$?+84+LB P c $D$?*84*X Q 03P?+ < 5X R 0333P?) 5X S 0333P? *$%5X T 0333P?&,((5X U 0333P?d*--5X V 0333P?)005X W 0333P?)<2\25X X 03ffP?):!^!5X Y 03ffP?)!!5X Z 03ffP?5##5X [ 03ffP?5""5X \ 03ffP?5C#g#5X ] 03ffP?)N"q"5X ^ 03ffP?52'V'5X _ 03ffP?5''5X ` 03ffP?)=&b&5X a 03ffP?)&&5X b 03ffP?)%%5X c 03ffP?*I%k%5X d 03ffP?+7)[)5X e 03ffP?&,))5X f 03ffP?-H*k*5X g 03ffP?/***5X h 03ffP?,W+|+5X i 03ffP?J++,5X j 03ffP?+h,,5X k 03ffP?E*7.Z.5X l 03ffP?5//5X m 03ffP?)G/j/5X n 03ffP?!+..5X o 03ffP?)11U15X p 03ffP?*225X q 03ffP?>*73Y35X r 03ffP?*335LB s c $D$?4(4LB t c $D$?R3(U3LB u c $D$?2(2LB v c $D$?0(0LB w c $D$?/(/LB x c $D$?j.(k.LB y c $D$?/-(2-LB z c $D$?+(+LB { c $D$?*(*LB | c $D$?)()! } B, ?5646 0 Z5Pk ( C ^y" ~ B ?P4C4 10 Z5Pk ( C ^y"  B  ?3C3 20 Z5Pk ( C ^y"  B(" ?1C2 30 Z5Pk ( C ^y"  B< ?0CM1 40 Z5Pk ( C ^y"  B? ?f/C0 50 Z5Pk ( C ^y"  BXH ?+.C. 60 Z5Pk ( C ^y"  BXc ?,C- 70 Z5Pk ( C ^y"  B`f ?+Ca, 80 Z5Pk ( C ^y"  Bn ?}*C*+ 90 Z5Pk ( C ^y#  B ?@)s) 100 Z5Pk ( C ^y*  H;茱 ?`6 7 25F0 Z5Pk ( C ^y2  H;h ?l6&!7 78F1"03333 Z5Pk ( C ^y2  H; ?Z68$%7 41F1"03333 Z5Pk ( C ^y2  H; ?o6C()7 36F1"03333 Z5Pk ( C ^y2  H;@ ?p6*-r.7 35F1"03333 Z5Pk ( C ^y2  H;@ֱ ?p600x17 82F1"03333 Z5Pk ( C ^y2  H;Hٱ ?s612 7 77F1"03333 Z5Pk ( C ^yL  c $$?m)F45LB  c $D$?4P44E  Hp_ж_ж ?,pX3 % of GFP positive cells"0ff Z5Pk ( C ^y08 (P7S3g8  P7(S3g8`   0333$?(7`*I8`   0333$?-7/8D   BH_ж_ж ?c*7-h8 Fonder (F0)    Z5Pk ( C ^y.   BL _ж_ж ?>/7P0h8 F1  Z5Pk ( C ^y`   03ff$?~0718.   B_ж_ж ?B2P7T388 F2  Z5Pk ( C ^yL  c $$?'5@Jp>1  BD%_ж_ж ?D8>D,9 Copy number/cell  Z5Pk ( C ^y!  B41 ?7 88?8 0 Z5Pk ( C ^y"  B= ?7:;?8 10 Z5Pk ( C ^y"  BH ?7=`>?8 20 Z5Pk ( C ^y"  BJ ?7i@7A?8 30 Z5Pk ( C ^y"  BdX ?78CD?8 40 Z5Pk ( C ^y"  Bq ?7EF?8 50 Z5Pk ( C ^y"  BW ?7HI?8 60 Z5Pk ( C ^yD  H|_ж_ж ?A*66v2 % of grenn lymphocytes"0ff Z5Pk ( C ^yj2  B$?(89')#  Bu ?(67]) 100 Z5Pk ( C ^y  @ABCE F *?(@.@8|85  @ABC-E Ff *?-@2.|88A0  @ABCE Ff *?)@A0@8|86  @ABC E Ff *? @)9:+*LB  c $D$?(48 I(X  0$?(48 I7LB  c $D$?(48687LB  c $D$?78487LB  c $D$?58485LB  c $D$?;4848=4LB  c $D$?p1848s1LB  c $D$?0-8481-LB  c $D$?+848+LB  c $D$?f*848g*LB  c $D$?(848(LB  c $D$?748 I7LB  c $D$?74868B7LB  c $D$?7;;B7LB  c $D$?7==B7LB  c $D$?7@@B7LB  c $D$?7qCtCB7LB  c $D$?7>F@FB7LB  c $D$?7 I IB7j2  B$?b68p86j2  B$?6w99z6j2  B$?/:H;W0j2  B$?)`;;B*j2  B$?*;[<+j2  B$?;4A::4j2  B$?648847j2  B$?)95:B*j2  B$?A)::)j2  B$?6p88z6j2  B$?59Y:?6j2  Bff$?d0(880j2  Bff$?G48|84j2  Bff$?28p82j2  Bff$?588b6j2  Bff$?68|847j2  Bff$?r+e88+j2  Bff$?04884j2  Bff$?828|82j2  Bff$?828|82j2  Bff$?0@881j2  Bff$?0L88o0j2  B$?_-X88-j2  B$?$48p84j2  B$?A08p80j2  B$?F38|83j2  B$?{0X880j2  B$?14881j2  B$?258p85j2  B$?`58p85j2  B$?-`;;-j2  B$?M*,<<*j2  B$?N14881j2  B$?68|847j2  B$?*==}*j2  B$?A)=;;)j2  B$?.;7,  @ABCEF3ff *? @F,,>>i,  @ABCE F3ff *?@",>>F,  @ABCEF3ff *? @+>?",  @ABCE F3ff *?@+?W?+  @ABCEF3ff *? @+W??+  @ABCE F3ff *?@+??+  @ABCE F3ff *?@+?.@+  @ABCEF3ff *? @~+.@@+  @ABCE F3ff *?@g+@@~+  @ABCEF3ff *? @P+@Ag+  @ABCE F3ff *?@8+AYAP+   @ABCEF3ff *? @ +YAA8+   @ABCE F3ff *?@+AA +   @ABCE F3ff *?@*A/B+   @ABCEF3ff *? @*/BB*   @ABCE F3ff *?@*BB*  @ABCEF3ff *? @*BC*  @ABCE F3ff *?@*CZC*  @ABCEF3ff *? @*ZCC*  @ABCE F3ff *?@*CC*  @ABCE F3ff *?@}*C1D*  @ABCEF3ff *? @q*1DD}*  @ABCE F3ff *?@f*DDq*  @ABCEF3ff *? @M*DEf*  @ABCE F3ff *?@B*E[EM*  @ABCEF3ff *? @+*[EEB*  @ABCE F3ff *?@*EE+*  @ABCE F3ff *?@*E2F*XB  0D3ff *?6P8X87  @ABC)E F333 *?)@2|88^4  @ABC#EF333 *?#  @0892  @ABCE F333 *?@)09S90  @ABCEF333 *?  @K/S99)0  @ABC E F333 *? @.99K/   @ABCE F333 *?@x.9:. ! @ABC EF333 *?  @-:}:x. " @ABCE F333 *?@-}::- # @ABC EF333 *?  @;-:;- $ @ABCE F333 *?@-;U;;- % @ABCEF333 *? @,U;;- & @ABCE F333 *?@t,;;, ' @ABCE F333 *?@F,;, + / @ABCEF333 *? @*,>>* 0 @ABCE F333 *?@*>>* 1 @ABCEF333 *? @*>?* 2 @ABCE F333 *?@q*?W?* 3 @ABCEF333 *? @M*W??q* 4 @ABCE F333 *?@7*??M* 5 @ABCE F333 *?@*?.@7* 6 @ABCEF333 *? @).@@* 7 @ABCE F333 *?@)@@) 8 @ABCEF333 *? @)@$A) 9 @ABCE F333 *?@)AYA) : @ABCEF333 *? @)YAA) ; @ABCE F333 *?@)AA) < @ABCE F333 *?@p)A/B) = @ABCEF333 *? @W)/BBp) > @ABCE F333 *?@A)BBW) ? @ABCEF333 *? @))BCA) @ @ABCE F333 *?@)CZC)) A @ABCEF333 *? @(ZCC) B @ABC'EFf *?'  @i,892. C @ABCE Ff *? @+9S9i, D @ABCEFf *?  @*]99+ E @ABC E Ff *? @+*99* F @ABC EFf *?  @)):}:) G # @ABC,E F *?,@,|88. H @ABC&EF *?&  @,+89, I @ABCE F *? @X*9S9,+ J @ABCEF *?  @p)S99X*! K B(ݸ ?66]7a7 0 Z5Pk ( C ^y" L B ?J5775 10 Z5Pk ( C ^y" M B ?4774 20 Z5Pk ( C ^y" N B ?27703 30 Z5Pk ( C ^y" O B ?B1771 40 Z5Pk ( C ^y" P B ?/77j0 50 Z5Pk ( C ^y" Q B" ?`.77 / 60 Z5Pk ( C ^y" R BX+ ?-77- 70 Z5Pk ( C ^y" S BXF ?+77L, 80 Z5Pk ( C ^y" T B`I ?*77* 90 Z5Pk ( C ^y0 U NQ_ж_ж3 ?*=?g+ SFFV Z5Pk ( C ^y< V Zl_ж_ж̙3f̙3f$?*@B+ FG12 Z5Pk ( C ^yB W Ny_ж_ж ?)8<* CAG (C57Bl6)  Z5Pk ( C ^y+ X N|_ж_жf ?*C9:+ CAG Z5Pk ( C ^yXB Y@ 0D3ff *?4k88?5XB Z@ 0D3ff *??5Q8o85R2 [ s * ?08G9B1j2 \ B$?"38 93XB ]@ 0D333 *?4k884XB ^@ 0D333 *?4Q8o85j2 _ B$?+ 9w9+LB ` c $D$?/828/LB a c $D$?.77.LB b c $D$?28282 c <_ж_ж ?9.6K+= TpFig. 4: GFP expression in Founders lymphocytes Correlation between GFP expression level in the lymphocytes and transgene copy number in the founders. CAG is the strongest promoter. T(f Z5Pk ( C ^y 8 `'@@ J d@`'@ Jb e <৺_ж_ж ?8A@ J 050-85% < 5 kb not limited multiple sites yes.) 1# 1Z5Pk ( C ^yY f <_ж_ж ?0A8 J )10% not limited limitided 1 site yes **# %Z5Pk ( C ^y g < ƺ_ж_ж ?`'A0 J R^% transgenic mice transgene size embryo background insertion site Germline transmission H# ;  ]##& #) Z5Pk ( C ^y4 h <˺_ж_ж ?8@@A Lentiviral vectors# Z5Pk ( C ^y3 i <h޺_ж_ж ?0@8A  DNA injection# Z5Pk ( C ^yT j c $ ?`'@0ATB k c $D81?`'A@ATB l c $Do?`' J@!JTB m c $Do?`'@a' JTB n c $D81?0@0 JTB o c $D81?8@8 JTB p c $Do?@@@ JTB q c $D81?0@8@TB r c $Do?`'@0@TB s c $Do?8@@@F t B_ж_ж ??-:"@ Comparison of the two technics"0ff Z5Pk ( C ^y/d8 P7G uP7G) v B_ж_ж ?@kB   Z5Pk ( C ^yT w c $ ?i89@ U7 : xU7 := y B _ж_ж ?#U7s!: vector ( " Z5Pk ( C ^y` z 0___$?U7!:@ U7& : {U7& :` | B_ж_ж ?U7!: *Viral titer (UI/ml) ( " Z5Pk ( C ^y` } 0___$?U7'!:@ 'U7 : ~'U7 :9  B0%_ж_ж ?IU7 !: Implanted embryos" Z5Pk ( C ^y`  0___$?'U7 !:@  U7 :  U7 :7  B 2_ж_ж ? U7 !: Newborn animals" Z5Pk ( C ^y`  0___$? U7!:@ U7: U7:@ V7t: V7t:d  <B_ж_ж ?!V7n:  Transgenic animals H  &" Z5Pk ( C ^yT  c $ ?T8u09`  0___$?U7!:@ U7 : U7 :?  BP_ж_ ?U7!:  Efficiency &     Z5Pk ( C ^y`  0___$?U7!:@ U73 : U73 :=  BV_ж_ж ?U7!: GFP expessing animals" Z5Pk ( C ^y`  0___$?U74!:@ 4U7 : 4U7 :3  B8j_ж_ж ?VU7h!:  Copy number  "  Z5Pk ( C ^y`  0___$?4U7!:@ !::< !::<O  <,x_ж_ ?#!:s;<  FG12-gfp <    &  Z5Pk ( C ^y`  0___$?!:;<@ !:&:< !:&:<H  <H_ж_ж ?!:;< 6.4 108 8 "*  Z5Pk ( C ^y`  0___$?!:';<@ '!: :< '!: :<"  <_ж_ж ?I!: ;< 93" Z5Pk ( C ^y`  0___$?'!: ;<@  !: :<  !: :<1  <_ж_ж ? !: ;< 23 & " Z5Pk ( C ^y`  0___$? !:;<@ !::< !::<=  <谻_ж_ж ?!!:p;< 17 2   & Z5Pk ( C ^y`  0___$?!:;<@ !::< !::<@  <h_ж_ж ?!:;< 74% 4   & Z5Pk ( C ^y`  0___$?!:;<@ !:3:< !:3:<3  <4̻_ж_ж ?!:;< 11 ( " Z5Pk ( C ^y`  0___$?!:4;<@ 4!::< 4!::<7  <ڻ_ж_ж ?V!:h;< 0.4-50 ( " Z5Pk ( C ^y`  0___$?4!:;<@ ;<= ;<=O  <4_ж_ж ?#;<s=  SFFV-gfp <    "&  Z5Pk ( C ^y`  0___$?;<=@ ;<&= ;<&=H  <(_ж_ж ?;<= 1.8 109 8 "*  Z5Pk ( C ^y`  0___$?;<'=@ ';< = ';< =2  <_ж_ж ?I;< = 102 & " Z5Pk ( C ^y`  0___$?';< =@  ;< =  ;< =1  <(_ж_ж ? ;< = 24 & " Z5Pk ( C ^y`  0___$? ;<=@ ;<= ;<==  <P"_ж_ж ?!;<p= 19 2   & Z5Pk ( C ^y`  0___$?;<=@ ;<= ;<=@  </_ж_ж ?;<= 80% 4   & Z5Pk ( C ^y`  0___$?;<=@ ;<3= ;<3=$  <<_ж_ж ?;<= 17" Z5Pk ( C ^y`  0___$?;<4=@ 4;<= 4;<=7  <s+G-Oh+'0xm PXx Prsentation PowerPoint mteixeirDJEBALI1@@ޅGl L6!4    """)))UUUMMMBBB999|PP3f333f3333f3ffffff3f̙3ff333f333333333f33333333f33f3ff3f3f3f3333f33̙33333f333333f3333f3ffffff3f33ff3f3f3f3fff3ffffffffff3ffff̙fff3fffff3fff333f3f3ff3ff33f̙̙3̙ff̙̙̙3f̙3f333f3333f3ffffff3f̙3f3f3f333f3333f3ffffff3f̙3f3ffffffffff!___www4!4A (dʦ """)))UUUMMMBBB999|PP3f3333f333ff3fffff3f3f̙f3333f3333333333f3333333f3f33ff3f3f3f3333f3333333f3̙33333f333ff3ffffff3f33f3ff3f3f3ffff3fffffffff3fffffff3f̙ffff3ff333f3ff33fff33f3ff̙3f3f3333f333ff3fffff̙̙3̙f̙̙̙3f̙3f3f3333f333ff3fffff3f3f̙3ffffffffff!___www'''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''qq,q'''''''q,,''2'''q'''q',q''2q'''q'''q''2q'''2q''2qq'''qq'''qq',qqq',2q'2222q'2qqqqqq'''''''''''''''''''''''''''''''''s#K's#K""C'''s#K'''qkp''ppJl#Jl'#Jl's#Kqqqq"p#Jqq'-'---------------------''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''''¼ÚSݼݼݼ'''ÙϮm˜❝Y+ݻܻܵ'™ݵϴmmmfmmmqqlwNrtݼݻݼݼmmmmm˜qqۺýü՜.+,0   Personnalis^ Times New RomanLucida Sans UnicodeVerdanaArialVerdana Greek Arial NarrowModle par dfautDiapositive 1 Polices utilisesModle de conceptionTitres des diapositives_g-DJEBALIDJEBALI  !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~      !"#$%&'()*+,-./0123456789:;<=>?@ABCDEFGHIJKLMNOPQRSTUVWXYZ[\]^_`abcdefghijklmnopqrstuvwxyz{|}~     Root EntrydO)PicturescCurrent UserSummaryInformation(mPowerPoint Document(-DocumentSummaryInformation8