﻿               SEQUENCE LISTING

<110> UNIVERSITÉ PIERRE ET MARIE CURIE - PARIS 6 (UPMC)
      APHP (Assistance Publique - Hôpitaux de Paris)
      INSTITUT NATIONAL DE LA SANTÉ ET DE LA RECHERCHE MÉDICALE
      (INSERM)
      HULOT Jean-Sébastien
      DUSSAUD Sébastien
      JEZIOROWSKA Dorota Monika

<120> PROCÉDÉ DE DIFFÉRENCIATION DE CELLULES SOUCHES PLURIPOTENTES EN CARDIOMYOCYTES

<130> CV - 713/PCT

<160> 24

<170> BiSSAP 1.3.6

<210> 1
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène MyoD1

<400> 1
gcctgggctc cggggcgttt                                                20


<210> 2
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène MyoD1

<400> 2
gggcccctgc ggccaccccg                                                20


<210> 3
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène MyoD1

<400> 3
gctccctccc tgcccggtag                                                20


<210> 4
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène MyoD1

<400> 4
gaggtttgga aagggcgtgc                                                20


<210> 5
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Ascl1

<400> 5
ggctgggtgt cccattgaaa                                                20


<210> 6
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Ascl1

<400> 6
gcagccgctc gctgcagcag                                                20


<210> 7
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Ascl1

<400> 7
gtggagagtt tgcaaggagc                                                20


<210> 8
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Ascl1

<400> 8
ggtttattca gccgggagtc                                                20


<210> 9
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Gata4

<400> 9
aacccaatcg acctccggct                                                20


<210> 10
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Gata4

<400> 10
gcccaggcaa gcctggggag                                                20


<210> 11
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Gata4

<400> 11
cacaggccag tcagcgccct                                                20


<210> 12
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Gata4

<400> 12
ctgaaggctg cggtaatcga                                                20


<210> 13
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Gata4

<400> 13
tagcatccag ccgggcaccc                                                20


<210> 14
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Mef2c

<400> 14
cgggggggat tgaaggatac                                                20


<210> 15
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Mef2c

<400> 15
cgactcgttt acgagtcctt                                                20


<210> 16
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Tbx5

<400> 16
aatcccccgc ggaggagctc                                                20


<210> 17
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Tbx5

<400> 17
gtggagggga cacgagacag                                                20


<210> 18
<211> 1368
<212> PRT
<213> Artificial Sequence


<220> 
<223> Cas9 dépourvue d’activité endonucléase dérivée de SP-Cas9

<400> 18
Met Asp Lys Lys Tyr Ser Ile Gly Leu Ala Ile Gly Thr Asn Ser Val 
1               5                   10                  15      
Gly Trp Ala Val Ile Thr Asp Glu Tyr Lys Val Pro Ser Lys Lys Phe 
            20                  25                  30          
Lys Val Leu Gly Asn Thr Asp Arg His Ser Ile Lys Lys Asn Leu Ile 
        35                  40                  45              
Gly Ala Leu Leu Phe Asp Ser Gly Glu Thr Ala Glu Ala Thr Arg Leu 
    50                  55                  60                  
Lys Arg Thr Ala Arg Arg Arg Tyr Thr Arg Arg Lys Asn Arg Ile Cys 
65                  70                  75                  80  
Tyr Leu Gln Glu Ile Phe Ser Asn Glu Met Ala Lys Val Asp Asp Ser 
                85                  90                  95      
Phe Phe His Arg Leu Glu Glu Ser Phe Leu Val Glu Glu Asp Lys Lys 
            100                 105                 110         
His Glu Arg His Pro Ile Phe Gly Asn Ile Val Asp Glu Val Ala Tyr 
        115                 120                 125             
His Glu Lys Tyr Pro Thr Ile Tyr His Leu Arg Lys Lys Leu Val Asp 
    130                 135                 140                 
Ser Thr Asp Lys Ala Asp Leu Arg Leu Ile Tyr Leu Ala Leu Ala His 
145                 150                 155                 160 
Met Ile Lys Phe Arg Gly His Phe Leu Ile Glu Gly Asp Leu Asn Pro 
                165                 170                 175     
Asp Asn Ser Asp Val Asp Lys Leu Phe Ile Gln Leu Val Gln Thr Tyr 
            180                 185                 190         
Asn Gln Leu Phe Glu Glu Asn Pro Ile Asn Ala Ser Gly Val Asp Ala 
        195                 200                 205             
Lys Ala Ile Leu Ser Ala Arg Leu Ser Lys Ser Arg Arg Leu Glu Asn 
    210                 215                 220                 
Leu Ile Ala Gln Leu Pro Gly Glu Lys Lys Asn Gly Leu Phe Gly Asn 
225                 230                 235                 240 
Leu Ile Ala Leu Ser Leu Gly Leu Thr Pro Asn Phe Lys Ser Asn Phe 
                245                 250                 255     
Asp Leu Ala Glu Asp Ala Lys Leu Gln Leu Ser Lys Asp Thr Tyr Asp 
            260                 265                 270         
Asp Asp Leu Asp Asn Leu Leu Ala Gln Ile Gly Asp Gln Tyr Ala Asp 
        275                 280                 285             
Leu Phe Leu Ala Ala Lys Asn Leu Ser Asp Ala Ile Leu Leu Ser Asp 
    290                 295                 300                 
Ile Leu Arg Val Asn Thr Glu Ile Thr Lys Ala Pro Leu Ser Ala Ser 
305                 310                 315                 320 
Met Ile Lys Arg Tyr Asp Glu His His Gln Asp Leu Thr Leu Leu Lys 
                325                 330                 335     
Ala Leu Val Arg Gln Gln Leu Pro Glu Lys Tyr Lys Glu Ile Phe Phe 
            340                 345                 350         
Asp Gln Ser Lys Asn Gly Tyr Ala Gly Tyr Ile Asp Gly Gly Ala Ser 
        355                 360                 365             
Gln Glu Glu Phe Tyr Lys Phe Ile Lys Pro Ile Leu Glu Lys Met Asp 
    370                 375                 380                 
Gly Thr Glu Glu Leu Leu Val Lys Leu Asn Arg Glu Asp Leu Leu Arg 
385                 390                 395                 400 
Lys Gln Arg Thr Phe Asp Asn Gly Ser Ile Pro His Gln Ile His Leu 
                405                 410                 415     
Gly Glu Leu His Ala Ile Leu Arg Arg Gln Glu Asp Phe Tyr Pro Phe 
            420                 425                 430         
Leu Lys Asp Asn Arg Glu Lys Ile Glu Lys Ile Leu Thr Phe Arg Ile 
        435                 440                 445             
Pro Tyr Tyr Val Gly Pro Leu Ala Arg Gly Asn Ser Arg Phe Ala Trp 
    450                 455                 460                 
Met Thr Arg Lys Ser Glu Glu Thr Ile Thr Pro Trp Asn Phe Glu Glu 
465                 470                 475                 480 
Val Val Asp Lys Gly Ala Ser Ala Gln Ser Phe Ile Glu Arg Met Thr 
                485                 490                 495     
Asn Phe Asp Lys Asn Leu Pro Asn Glu Lys Val Leu Pro Lys His Ser 
            500                 505                 510         
Leu Leu Tyr Glu Tyr Phe Thr Val Tyr Asn Glu Leu Thr Lys Val Lys 
        515                 520                 525             
Tyr Val Thr Glu Gly Met Arg Lys Pro Ala Phe Leu Ser Gly Glu Gln 
    530                 535                 540                 
Lys Lys Ala Ile Val Asp Leu Leu Phe Lys Thr Asn Arg Lys Val Thr 
545                 550                 555                 560 
Val Lys Gln Leu Lys Glu Asp Tyr Phe Lys Lys Ile Glu Cys Phe Asp 
                565                 570                 575     
Ser Val Glu Ile Ser Gly Val Glu Asp Arg Phe Asn Ala Ser Leu Gly 
            580                 585                 590         
Thr Tyr His Asp Leu Leu Lys Ile Ile Lys Asp Lys Asp Phe Leu Asp 
        595                 600                 605             
Asn Glu Glu Asn Glu Asp Ile Leu Glu Asp Ile Val Leu Thr Leu Thr 
    610                 615                 620                 
Leu Phe Glu Asp Arg Glu Met Ile Glu Glu Arg Leu Lys Thr Tyr Ala 
625                 630                 635                 640 
His Leu Phe Asp Asp Lys Val Met Lys Gln Leu Lys Arg Arg Arg Tyr 
                645                 650                 655     
Thr Gly Trp Gly Arg Leu Ser Arg Lys Leu Ile Asn Gly Ile Arg Asp 
            660                 665                 670         
Lys Gln Ser Gly Lys Thr Ile Leu Asp Phe Leu Lys Ser Asp Gly Phe 
        675                 680                 685             
Ala Asn Arg Asn Phe Met Gln Leu Ile His Asp Asp Ser Leu Thr Phe 
    690                 695                 700                 
Lys Glu Asp Ile Gln Lys Ala Gln Val Ser Gly Gln Gly Asp Ser Leu 
705                 710                 715                 720 
His Glu His Ile Ala Asn Leu Ala Gly Ser Pro Ala Ile Lys Lys Gly 
                725                 730                 735     
Ile Leu Gln Thr Val Lys Val Val Asp Glu Leu Val Lys Val Met Gly 
            740                 745                 750         
Arg His Lys Pro Glu Asn Ile Val Ile Glu Met Ala Arg Glu Asn Gln 
        755                 760                 765             
Thr Thr Gln Lys Gly Gln Lys Asn Ser Arg Glu Arg Met Lys Arg Ile 
    770                 775                 780                 
Glu Glu Gly Ile Lys Glu Leu Gly Ser Gln Ile Leu Lys Glu His Pro 
785                 790                 795                 800 
Val Glu Asn Thr Gln Leu Gln Asn Glu Lys Leu Tyr Leu Tyr Tyr Leu 
                805                 810                 815     
Gln Asn Gly Arg Asp Met Tyr Val Asp Gln Glu Leu Asp Ile Asn Arg 
            820                 825                 830         
Leu Ser Asp Tyr Asp Val Ala Ala Ile Val Pro Gln Ser Phe Leu Lys 
        835                 840                 845             
Asp Asp Ser Ile Asp Asn Lys Val Leu Thr Arg Ser Asp Lys Ala Arg 
    850                 855                 860                 
Gly Lys Ser Asp Asn Val Pro Ser Glu Glu Val Val Lys Lys Met Lys 
865                 870                 875                 880 
Asn Tyr Trp Arg Gln Leu Leu Asn Ala Lys Leu Ile Thr Gln Arg Lys 
                885                 890                 895     
Phe Asp Asn Leu Thr Lys Ala Glu Arg Gly Gly Leu Ser Glu Leu Asp 
            900                 905                 910         
Lys Ala Gly Phe Ile Lys Arg Gln Leu Val Glu Thr Arg Gln Ile Thr 
        915                 920                 925             
Lys His Val Ala Gln Ile Leu Asp Ser Arg Met Asn Thr Lys Tyr Asp 
    930                 935                 940                 
Glu Asn Asp Lys Leu Ile Arg Glu Val Lys Val Ile Thr Leu Lys Ser 
945                 950                 955                 960 
Lys Leu Val Ser Asp Phe Arg Lys Asp Phe Gln Phe Tyr Lys Val Arg 
                965                 970                 975     
Glu Ile Asn Asn Tyr His His Ala His Asp Ala Tyr Leu Asn Ala Val 
            980                 985                 990         
Val Gly Thr Ala Leu Ile Lys Lys Tyr Pro Lys Leu Glu Ser Glu Phe 
        995                 1000                1005            
Val Tyr Gly Asp Tyr Lys Val Tyr Asp Val Arg Lys Met Ile Ala Lys 
    1010                1015                1020                
Ser Glu Gln Glu Ile Gly Lys Ala Thr Ala Lys Tyr Phe Phe Tyr Ser 
1025                1030                1035                1040
Asn Ile Met Asn Phe Phe Lys Thr Glu Ile Thr Leu Ala Asn Gly Glu 
                1045                1050                1055    
Ile Arg Lys Arg Pro Leu Ile Glu Thr Asn Gly Glu Thr Gly Glu Ile 
            1060                1065                1070        
Val Trp Asp Lys Gly Arg Asp Phe Ala Thr Val Arg Lys Val Leu Ser 
        1075                1080                1085            
Met Pro Gln Val Asn Ile Val Lys Lys Thr Glu Val Gln Thr Gly Gly 
    1090                1095                1100                
Phe Ser Lys Glu Ser Ile Leu Pro Lys Arg Asn Ser Asp Lys Leu Ile 
1105                1110                1115                1120
Ala Arg Lys Lys Asp Trp Asp Pro Lys Lys Tyr Gly Gly Phe Asp Ser 
                1125                1130                1135    
Pro Thr Val Ala Tyr Ser Val Leu Val Val Ala Lys Val Glu Lys Gly 
            1140                1145                1150        
Lys Ser Lys Lys Leu Lys Ser Val Lys Glu Leu Leu Gly Ile Thr Ile 
        1155                1160                1165            
Met Glu Arg Ser Ser Phe Glu Lys Asn Pro Ile Asp Phe Leu Glu Ala 
    1170                1175                1180                
Lys Gly Tyr Lys Glu Val Lys Lys Asp Leu Ile Ile Lys Leu Pro Lys 
1185                1190                1195                1200
Tyr Ser Leu Phe Glu Leu Glu Asn Gly Arg Lys Arg Met Leu Ala Ser 
                1205                1210                1215    
Ala Gly Glu Leu Gln Lys Gly Asn Glu Leu Ala Leu Pro Ser Lys Tyr 
            1220                1225                1230        
Val Asn Phe Leu Tyr Leu Ala Ser His Tyr Glu Lys Leu Lys Gly Ser 
        1235                1240                1245            
Pro Glu Asp Asn Glu Gln Lys Gln Leu Phe Val Glu Gln His Lys His 
    1250                1255                1260                
Tyr Leu Asp Glu Ile Ile Glu Gln Ile Ser Glu Phe Ser Lys Arg Val 
1265                1270                1275                1280
Ile Leu Ala Asp Ala Asn Leu Asp Lys Val Leu Ser Ala Tyr Asn Lys 
                1285                1290                1295    
His Arg Asp Lys Pro Ile Arg Glu Gln Ala Glu Asn Ile Ile His Leu 
            1300                1305                1310        
Phe Thr Leu Thr Asn Leu Gly Ala Pro Ala Ala Phe Lys Tyr Phe Asp 
        1315                1320                1325            
Thr Thr Ile Asp Arg Lys Arg Tyr Thr Ser Thr Lys Glu Val Leu Asp 
    1330                1335                1340                
Ala Thr Leu Ile His Gln Ser Ile Thr Gly Leu Tyr Glu Thr Arg Ile 
1345                1350                1355                1360
Asp Leu Ser Gln Leu Gly Gly Asp 
                1365            

<210> 19
<211> 11
<212> PRT
<213> Artificial Sequence


<220> 
<223> VP16

<400> 19
Asp Ala Leu Asp Asp Phe Asp Leu Asp Met Leu 
1               5                   10      

<210> 20
<211> 50
<212> PRT
<213> Artificial Sequence


<220> 
<223> VP64

<400> 20
Asp Ala Leu Asp Asp Phe Asp Leu Asp Met Leu Gly Ser Asp Ala Leu 
1               5                   10                  15      
Asp Asp Phe Asp Leu Asp Met Leu Gly Ser Asp Ala Leu Asp Asp Phe 
            20                  25                  30          
Asp Leu Asp Met Leu Gly Ser Asp Ala Leu Asp Asp Phe Asp Leu Asp 
        35                  40                  45              
Met Leu 
    50  

<210> 21
<211> 260
<212> PRT
<213> Artificial Sequence


<220> 
<223> p65

<400> 21
Gln Tyr Leu Pro Asp Thr Asp Asp Arg His Arg Ile Glu Glu Lys Arg 
1               5                   10                  15      
Lys Arg Thr Tyr Glu Thr Phe Lys Ser Ile Met Lys Lys Ser Pro Phe 
            20                  25                  30          
Ser Gly Pro Thr Asp Pro Arg Pro Pro Pro Arg Arg Ile Ala Val Pro 
        35                  40                  45              
Ser Arg Ser Ser Ala Ser Val Pro Lys Pro Ala Pro Gln Pro Tyr Pro 
    50                  55                  60                  
Phe Thr Ser Ser Leu Ser Thr Ile Asn Tyr Asp Glu Phe Pro Thr Met 
65                  70                  75                  80  
Val Phe Pro Ser Gly Gln Ile Ser Gln Ala Ser Ala Leu Ala Pro Ala 
                85                  90                  95      
Pro Pro Gln Val Leu Pro Gln Ala Pro Ala Pro Ala Pro Ala Pro Ala 
            100                 105                 110         
Met Val Ser Ala Leu Ala Gln Ala Pro Ala Pro Val Pro Val Leu Ala 
        115                 120                 125             
Pro Gly Pro Pro Gln Ala Val Ala Pro Pro Ala Pro Lys Pro Thr Gln 
    130                 135                 140                 
Ala Gly Glu Gly Thr Leu Ser Glu Ala Leu Leu Gln Leu Gln Phe Asp 
145                 150                 155                 160 
Asp Glu Asp Leu Gly Ala Leu Leu Gly Asn Ser Thr Asp Pro Ala Val 
                165                 170                 175     
Phe Thr Asp Leu Ala Ser Val Asp Asn Ser Glu Phe Gln Gln Leu Leu 
            180                 185                 190         
Asn Gln Gly Ile Pro Val Ala Pro His Thr Thr Glu Pro Met Leu Met 
        195                 200                 205             
Glu Tyr Pro Glu Ala Ile Thr Arg Leu Val Thr Gly Ala Gln Arg Pro 
    210                 215                 220                 
Pro Asp Pro Ala Pro Ala Pro Leu Gly Ala Pro Gly Leu Pro Asn Gly 
225                 230                 235                 240 
Leu Leu Ser Gly Asp Glu Asp Phe Ser Ser Ile Ala Asp Met Asp Phe 
                245                 250                 255     
Ser Ala Leu Leu 
            260 

<210> 22
<211> 190
<212> PRT
<213> Artificial Sequence


<220> 
<223> Rta

<400> 22
Arg Asp Ser Arg Glu Gly Met Phe Leu Pro Lys Pro Glu Ala Gly Ser 
1               5                   10                  15      
Ala Ile Ser Asp Val Phe Glu Gly Arg Glu Val Cys Gln Pro Lys Arg 
            20                  25                  30          
Ile Arg Pro Phe His Pro Pro Gly Ser Pro Trp Ala Asn Arg Pro Leu 
        35                  40                  45              
Pro Ala Ser Leu Ala Pro Thr Pro Thr Gly Pro Val His Glu Pro Val 
    50                  55                  60                  
Gly Ser Leu Thr Pro Ala Pro Val Pro Gln Pro Leu Asp Pro Ala Pro 
65                  70                  75                  80  
Ala Val Thr Pro Glu Ala Ser His Leu Leu Glu Asp Pro Asp Glu Glu 
                85                  90                  95      
Thr Ser Gln Ala Val Lys Ala Leu Arg Glu Met Ala Asp Thr Val Ile 
            100                 105                 110         
Pro Gln Lys Glu Glu Ala Ala Ile Cys Gly Gln Met Asp Leu Ser His 
        115                 120                 125             
Pro Pro Pro Arg Gly His Leu Asp Glu Leu Thr Thr Thr Leu Glu Ser 
    130                 135                 140                 
Met Thr Glu Asp Leu Asn Leu Asp Ser Pro Leu Thr Pro Glu Leu Asn 
145                 150                 155                 160 
Glu Ile Leu Asp Thr Phe Leu Asn Asp Glu Cys Leu Leu His Ala Met 
                165                 170                 175     
His Ile Ser Thr Gly Leu Ser Ile Phe Asp Thr Ser Leu Phe 
            180                 185                 190 


<210> 23
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Mef2c

<400> 23
gatgatttct ccagctcgcg                                                20


<210> 24
<211> 20
<212> DNA
<213> Homo sapiens


<220> 
<223> cible de l'ARNgs pour le gène Mef2c

<400> 24
tctccgagcg tcctccgcag                                                20