                         SEQUENCE LISTING

<110>  JUNO THERAPEUTICS, INC.
       Odegard, Valerie
 
<120>  COMPOSITION AND METHODS FOR REGULATING INHIBITORY INTERACTIONS IN
        GENETICALLY ENGINEERED CELL

<130>  735042002440

<140>  Not Yet Assigned
<141>  Concurrently Herewith

<150>  62/168,721
<151>  2015-05-29

<150>  62/244,132
<151>  2015-10-20

<160>  24    

<170>  PatentIn version 3.5

<210>  1
<211>  25
<212>  RNA
<213>  Artificial Sequence

<220>
<223>  siRNA specific for PD-L1

<400>  1
aaugcguuca gcaaaugcca guagg                                             25


<210>  2
<211>  19
<212>  RNA
<213>  Artificial Sequence

<220>
<223>  siRNA specific for PD-L1

<400>  2
cuaauugucu auugggaaa                                                    19


<210>  3
<211>  19
<212>  RNA
<213>  Artificial Sequence

<220>
<223>  siRNA specific for PD-L1

<400>  3
cgacuacaag cgaauuacu                                                    19


<210>  4
<211>  25
<212>  RNA
<213>  Artificial Sequence

<220>
<223>  siRNA specific for PD-L1 (sense sequence)

<400>  4
ccuacuggca uuugcugaac gcauu                                             25


<210>  5
<211>  25
<212>  RNA
<213>  Artificial Sequence

<220>
<223>  siRNA specific for PD-L1 (anti-sense sequence)

<400>  5
aaugcguuca gcaaaugcca guagg                                             25


<210>  6
<211>  25
<212>  RNA
<213>  Artificial Sequence

<220>
<223>  siRNA specific for PD-1

<400>  6
uuacgucucc uccaaaugug uauca                                             25


<210>  7
<211>  290
<212>  PRT
<213>  Homo sapiens


<220>
<221>  MISC_FEATURE
<223>  PD-L1

<400>  7

Met Arg Ile Phe Ala Val Phe Ile Phe Met Thr Tyr Trp His Leu Leu 
1               5                   10                  15      


Asn Ala Phe Thr Val Thr Val Pro Lys Asp Leu Tyr Val Val Glu Tyr 
            20                  25                  30          


Gly Ser Asn Met Thr Ile Glu Cys Lys Phe Pro Val Glu Lys Gln Leu 
        35                  40                  45              


Asp Leu Ala Ala Leu Ile Val Tyr Trp Glu Met Glu Asp Lys Asn Ile 
    50                  55                  60                  


Ile Gln Phe Val His Gly Glu Glu Asp Leu Lys Val Gln His Ser Ser 
65                  70                  75                  80  


Tyr Arg Gln Arg Ala Arg Leu Leu Lys Asp Gln Leu Ser Leu Gly Asn 
                85                  90                  95      


Ala Ala Leu Gln Ile Thr Asp Val Lys Leu Gln Asp Ala Gly Val Tyr 
            100                 105                 110         


Arg Cys Met Ile Ser Tyr Gly Gly Ala Asp Tyr Lys Arg Ile Thr Val 
        115                 120                 125             


Lys Val Asn Ala Pro Tyr Asn Lys Ile Asn Gln Arg Ile Leu Val Val 
    130                 135                 140                 


Asp Pro Val Thr Ser Glu His Glu Leu Thr Cys Gln Ala Glu Gly Tyr 
145                 150                 155                 160 


Pro Lys Ala Glu Val Ile Trp Thr Ser Ser Asp His Gln Val Leu Ser 
                165                 170                 175     


Gly Lys Thr Thr Thr Thr Asn Ser Lys Arg Glu Glu Lys Leu Phe Asn 
            180                 185                 190         


Val Thr Ser Thr Leu Arg Ile Asn Thr Thr Thr Asn Glu Ile Phe Tyr 
        195                 200                 205             


Cys Thr Phe Arg Arg Leu Asp Pro Glu Glu Asn His Thr Ala Glu Leu 
    210                 215                 220                 


Val Ile Pro Glu Leu Pro Leu Ala His Pro Pro Asn Glu Arg Thr His 
225                 230                 235                 240 


Leu Val Ile Leu Gly Ala Ile Leu Leu Cys Leu Gly Val Ala Leu Thr 
                245                 250                 255     


Phe Ile Phe Arg Leu Arg Lys Gly Arg Met Met Asp Val Lys Lys Cys 
            260                 265                 270         


Gly Ile Gln Asp Thr Asn Ser Lys Lys Gln Ser Asp Thr His Leu Glu 
        275                 280                 285             


Glu Thr 
    290 


<210>  8
<211>  3691
<212>  DNA
<213>  Homo sapiens


<220>
<221>  misc_feature
<223>  CD274 encoding PD-L1

<400>  8
ggcgcaacgc tgagcagctg gcgcgtcccg cgcggcccca gttctgcgca gcttcccgag       60

gctccgcacc agccgcgctt ctgtccgcct gcagggcatt ccagaaagat gaggatattt      120

gctgtcttta tattcatgac ctactggcat ttgctgaacg catttactgt cacggttccc      180

aaggacctat atgtggtaga gtatggtagc aatatgacaa ttgaatgcaa attcccagta      240

gaaaaacaat tagacctggc tgcactaatt gtctattggg aaatggagga taagaacatt      300

attcaatttg tgcatggaga ggaagacctg aaggttcagc atagtagcta cagacagagg      360

gcccggctgt tgaaggacca gctctccctg ggaaatgctg cacttcagat cacagatgtg      420

aaattgcagg atgcaggggt gtaccgctgc atgatcagct atggtggtgc cgactacaag      480

cgaattactg tgaaagtcaa tgccccatac aacaaaatca accaaagaat tttggttgtg      540

gatccagtca cctctgaaca tgaactgaca tgtcaggctg agggctaccc caaggccgaa      600

gtcatctgga caagcagtga ccatcaagtc ctgagtggta agaccaccac caccaattcc      660

aagagagagg agaagctttt caatgtgacc agcacactga gaatcaacac aacaactaat      720

gagattttct actgcacttt taggagatta gatcctgagg aaaaccatac agctgaattg      780

gtcatcccag aactacctct ggcacatcct ccaaatgaaa ggactcactt ggtaattctg      840

ggagccatct tattatgcct tggtgtagca ctgacattca tcttccgttt aagaaaaggg      900

agaatgatgg atgtgaaaaa atgtggcatc caagatacaa actcaaagaa gcaaagtgat      960

acacatttgg aggagacgta atccagcatt ggaacttctg atcttcaagc agggattctc     1020

aacctgtggt ttaggggttc atcggggctg agcgtgacaa gaggaaggaa tgggcccgtg     1080

ggatgcaggc aatgtgggac ttaaaaggcc caagcactga aaatggaacc tggcgaaagc     1140

agaggaggag aatgaagaaa gatggagtca aacagggagc ctggagggag accttgatac     1200

tttcaaatgc ctgaggggct catcgacgcc tgtgacaggg agaaaggata cttctgaaca     1260

aggagcctcc aagcaaatca tccattgctc atcctaggaa gacgggttga gaatccctaa     1320

tttgagggtc agttcctgca gaagtgccct ttgcctccac tcaatgcctc aatttgtttt     1380

ctgcatgact gagagtctca gtgttggaac gggacagtat ttatgtatga gtttttccta     1440

tttattttga gtctgtgagg tcttcttgtc atgtgagtgt ggttgtgaat gatttctttt     1500

gaagatatat tgtagtagat gttacaattt tgtcgccaaa ctaaacttgc tgcttaatga     1560

tttgctcaca tctagtaaaa catggagtat ttgtaaggtg cttggtctcc tctataacta     1620

caagtataca ttggaagcat aaagatcaaa ccgttggttg cataggatgt cacctttatt     1680

taacccatta atactctggt tgacctaatc ttattctcag acctcaagtg tctgtgcagt     1740

atctgttcca tttaaatatc agctttacaa ttatgtggta gcctacacac ataatctcat     1800

ttcatcgctg taaccaccct gttgtgataa ccactattat tttacccatc gtacagctga     1860

ggaagcaaac agattaagta acttgcccaa accagtaaat agcagacctc agactgccac     1920

ccactgtcct tttataatac aatttacagc tatattttac tttaagcaat tcttttattc     1980

aaaaaccatt tattaagtgc ccttgcaata tcaatcgctg tgccaggcat tgaatctaca     2040

gatgtgagca agacaaagta cctgtcctca aggagctcat agtataatga ggagattaac     2100

aagaaaatgt attattacaa tttagtccag tgtcatagca taaggatgat gcgaggggaa     2160

aacccgagca gtgttgccaa gaggaggaaa taggccaatg tggtctggga cggttggata     2220

tacttaaaca tcttaataat cagagtaatt ttcatttaca aagagaggtc ggtacttaaa     2280

ataaccctga aaaataacac tggaattcct tttctagcat tatatttatt cctgatttgc     2340

ctttgccata taatctaatg cttgtttata tagtgtctgg tattgtttaa cagttctgtc     2400

ttttctattt aaatgccact aaattttaaa ttcatacctt tccatgattc aaaattcaaa     2460

agatcccatg ggagatggtt ggaaaatctc cacttcatcc tccaagccat tcaagtttcc     2520

tttccagaag caactgctac tgcctttcat tcatatgttc ttctaaagat agtctacatt     2580

tggaaatgta tgttaaaagc acgtattttt aaaatttttt tcctaaatag taacacattg     2640

tatgtctgct gtgtactttg ctatttttat ttattttagt gtttcttata tagcagatgg     2700

aatgaatttg aagttcccag ggctgaggat ccatgccttc tttgtttcta agttatcttt     2760

cccatagctt ttcattatct ttcatatgat ccagtatatg ttaaatatgt cctacatata     2820

catttagaca accaccattt gttaagtatt tgctctagga cagagtttgg atttgtttat     2880

gtttgctcaa aaggagaccc atgggctctc cagggtgcac tgagtcaatc tagtcctaaa     2940

aagcaatctt attattaact ctgtatgaca gaatcatgtc tggaactttt gttttctgct     3000

ttctgtcaag tataaacttc actttgatgc tgtacttgca aaatcacatt ttctttctgg     3060

aaattccggc agtgtacctt gactgctagc taccctgtgc cagaaaagcc tcattcgttg     3120

tgcttgaacc cttgaatgcc accagctgtc atcactacac agccctccta agaggcttcc     3180

tggaggtttc gagattcaga tgccctggga gatcccagag tttcctttcc ctcttggcca     3240

tattctggtg tcaatgacaa ggagtacctt ggctttgcca catgtcaagg ctgaagaaac     3300

agtgtctcca acagagctcc ttgtgttatc tgtttgtaca tgtgcatttg tacagtaatt     3360

ggtgtgacag tgttctttgt gtgaattaca ggcaagaatt gtggctgagc aaggcacata     3420

gtctactcag tctattccta agtcctaact cctccttgtg gtgttggatt tgtaaggcac     3480

tttatccctt ttgtctcatg tttcatcgta aatggcatag gcagagatga tacctaattc     3540

tgcatttgat tgtcactttt tgtacctgca ttaatttaat aaaatattct tatttatttt     3600

gttacttggt acaccagcat gtccattttc ttgtttattt tgtgtttaat aaaatgttca     3660

gtttaacatc ccagtggaga aagttaaaaa a                                    3691


<210>  9
<211>  288
<212>  PRT
<213>  Homo sapiens


<220>
<221>  MISC_FEATURE
<223>  PD-1

<400>  9

Met Gln Ile Pro Gln Ala Pro Trp Pro Val Val Trp Ala Val Leu Gln 
1               5                   10                  15      


Leu Gly Trp Arg Pro Gly Trp Phe Leu Asp Ser Pro Asp Arg Pro Trp 
            20                  25                  30          


Asn Pro Pro Thr Phe Ser Pro Ala Leu Leu Val Val Thr Glu Gly Asp 
        35                  40                  45              


Asn Ala Thr Phe Thr Cys Ser Phe Ser Asn Thr Ser Glu Ser Phe Val 
    50                  55                  60                  


Leu Asn Trp Tyr Arg Met Ser Pro Ser Asn Gln Thr Asp Lys Leu Ala 
65                  70                  75                  80  


Ala Phe Pro Glu Asp Arg Ser Gln Pro Gly Gln Asp Cys Arg Phe Arg 
                85                  90                  95      


Val Thr Gln Leu Pro Asn Gly Arg Asp Phe His Met Ser Val Val Arg 
            100                 105                 110         


Ala Arg Arg Asn Asp Ser Gly Thr Tyr Leu Cys Gly Ala Ile Ser Leu 
        115                 120                 125             


Ala Pro Lys Ala Gln Ile Lys Glu Ser Leu Arg Ala Glu Leu Arg Val 
    130                 135                 140                 


Thr Glu Arg Arg Ala Glu Val Pro Thr Ala His Pro Ser Pro Ser Pro 
145                 150                 155                 160 


Arg Pro Ala Gly Gln Phe Gln Thr Leu Val Val Gly Val Val Gly Gly 
                165                 170                 175     


Leu Leu Gly Ser Leu Val Leu Leu Val Trp Val Leu Ala Val Ile Cys 
            180                 185                 190         


Ser Arg Ala Ala Arg Gly Thr Ile Gly Ala Arg Arg Thr Gly Gln Pro 
        195                 200                 205             


Leu Lys Glu Asp Pro Ser Ala Val Pro Val Phe Ser Val Asp Tyr Gly 
    210                 215                 220                 


Glu Leu Asp Phe Gln Trp Arg Glu Lys Thr Pro Glu Pro Pro Val Pro 
225                 230                 235                 240 


Cys Val Pro Glu Gln Thr Glu Tyr Ala Thr Ile Val Phe Pro Ser Gly 
                245                 250                 255     


Met Gly Thr Ser Ser Pro Ala Arg Arg Gly Ser Ala Asp Gly Pro Arg 
            260                 265                 270         


Ser Ala Gln Pro Leu Arg Pro Glu Asp Gly His Cys Ser Trp Pro Leu 
        275                 280                 285             


<210>  10
<211>  2115
<212>  DNA
<213>  Homo sapiens


<220>
<221>  misc_feature
<223>  PDCD1 encoding PD-1

<400>  10
agtttccctt ccgctcacct ccgcctgagc agtggagaag gcggcactct ggtggggctg       60

ctccaggcat gcagatccca caggcgccct ggccagtcgt ctgggcggtg ctacaactgg      120

gctggcggcc aggatggttc ttagactccc cagacaggcc ctggaacccc cccaccttct      180

ccccagccct gctcgtggtg accgaagggg acaacgccac cttcacctgc agcttctcca      240

acacatcgga gagcttcgtg ctaaactggt accgcatgag ccccagcaac cagacggaca      300

agctggccgc cttccccgag gaccgcagcc agcccggcca ggactgccgc ttccgtgtca      360

cacaactgcc caacgggcgt gacttccaca tgagcgtggt cagggcccgg cgcaatgaca      420

gcggcaccta cctctgtggg gccatctccc tggcccccaa ggcgcagatc aaagagagcc      480

tgcgggcaga gctcagggtg acagagagaa gggcagaagt gcccacagcc caccccagcc      540

cctcacccag gccagccggc cagttccaaa ccctggtggt tggtgtcgtg ggcggcctgc      600

tgggcagcct ggtgctgcta gtctgggtcc tggccgtcat ctgctcccgg gccgcacgag      660

ggacaatagg agccaggcgc accggccagc ccctgaagga ggacccctca gccgtgcctg      720

tgttctctgt ggactatggg gagctggatt tccagtggcg agagaagacc ccggagcccc      780

ccgtgccctg tgtccctgag cagacggagt atgccaccat tgtctttcct agcggaatgg      840

gcacctcatc ccccgcccgc aggggctcag ctgacggccc tcggagtgcc cagccactga      900

ggcctgagga tggacactgc tcttggcccc tctgaccggc ttccttggcc accagtgttc      960

tgcagaccct ccaccatgag cccgggtcag cgcatttcct caggagaagc aggcagggtg     1020

caggccattg caggccgtcc aggggctgag ctgcctgggg gcgaccgggg ctccagcctg     1080

cacctgcacc aggcacagcc ccaccacagg actcatgtct caatgcccac agtgagccca     1140

ggcagcaggt gtcaccgtcc cctacaggga gggccagatg cagtcactgc ttcaggtcct     1200

gccagcacag agctgcctgc gtccagctcc ctgaatctct gctgctgctg ctgctgctgc     1260

tgctgctgcc tgcggcccgg ggctgaaggc gccgtggccc tgcctgacgc cccggagcct     1320

cctgcctgaa cttgggggct ggttggagat ggccttggag cagccaaggt gcccctggca     1380

gtggcatccc gaaacgccct ggacgcaggg cccaagactg ggcacaggag tgggaggtac     1440

atggggctgg ggactcccca ggagttatct gctccctgca ggcctagaga agtttcaggg     1500

aaggtcagaa gagctcctgg ctgtggtggg cagggcagga aacccctcca cctttacaca     1560

tgcccaggca gcacctcagg ccctttgtgg ggcagggaag ctgaggcagt aagcgggcag     1620

gcagagctgg aggcctttca ggcccagcca gcactctggc ctcctgccgc cgcattccac     1680

cccagcccct cacaccactc gggagaggga catcctacgg tcccaaggtc aggagggcag     1740

ggctggggtt gactcaggcc cctcccagct gtggccacct gggtgttggg agggcagaag     1800

tgcaggcacc tagggccccc catgtgccca ccctgggagc tctccttgga acccattcct     1860

gaaattattt aaaggggttg gccgggctcc caccagggcc tgggtgggaa ggtacaggcg     1920

ttcccccggg gcctagtacc cccgccgtgg cctatccact cctcacatcc acacactgca     1980

cccccactcc tggggcaggg ccaccagcat ccaggcggcc agcaggcacc tgagtggctg     2040

ggacaaggga tcccccttcc ctgtggttct attatattat aattataatt aaatatgaga     2100

gcatgctaag gaaaa                                                      2115


<210>  11
<211>  1368
<212>  PRT
<213>  Streptococcus pyogenes


<220>
<221>  MISC_FEATURE
<223>  S. Pyogenes Cas9 Q99ZW2

<400>  11

Met Asp Lys Lys Tyr Ser Ile Gly Leu Asp 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 Asp His 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 Asn 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 
    1010                 1015                 1020             


Lys Ser  Glu Gln Glu Ile Gly  Lys Ala Thr Ala Lys  Tyr Phe Phe 
    1025                 1030                 1035             


Tyr Ser  Asn Ile Met Asn Phe  Phe Lys Thr Glu Ile  Thr Leu Ala 
    1040                 1045                 1050             


Asn Gly  Glu Ile Arg Lys Arg  Pro Leu Ile Glu Thr  Asn Gly Glu 
    1055                 1060                 1065             


Thr Gly  Glu Ile Val Trp Asp  Lys Gly Arg Asp Phe  Ala Thr Val 
    1070                 1075                 1080             


Arg Lys  Val Leu Ser Met Pro  Gln Val Asn Ile Val  Lys Lys Thr 
    1085                 1090                 1095             


Glu Val  Gln Thr Gly Gly Phe  Ser Lys Glu Ser Ile  Leu Pro Lys 
    1100                 1105                 1110             


Arg Asn  Ser Asp Lys Leu Ile  Ala Arg Lys Lys Asp  Trp Asp Pro 
    1115                 1120                 1125             


Lys Lys  Tyr Gly Gly Phe Asp  Ser Pro Thr Val Ala  Tyr Ser Val 
    1130                 1135                 1140             


Leu Val  Val Ala Lys Val Glu  Lys Gly Lys Ser Lys  Lys Leu Lys 
    1145                 1150                 1155             


Ser Val  Lys Glu Leu Leu Gly  Ile Thr Ile Met Glu  Arg Ser Ser 
    1160                 1165                 1170             


Phe Glu  Lys Asn Pro Ile Asp  Phe Leu Glu Ala Lys  Gly Tyr Lys 
    1175                 1180                 1185             


Glu Val  Lys Lys Asp Leu Ile  Ile Lys Leu Pro Lys  Tyr Ser Leu 
    1190                 1195                 1200             


Phe Glu  Leu Glu Asn Gly Arg  Lys Arg Met Leu Ala  Ser Ala Gly 
    1205                 1210                 1215             


Glu Leu  Gln Lys Gly Asn Glu  Leu Ala Leu Pro Ser  Lys Tyr Val 
    1220                 1225                 1230             


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 
    1250                 1255                 1260             


His Tyr  Leu Asp Glu Ile Ile  Glu Gln Ile Ser Glu  Phe Ser Lys 
    1265                 1270                 1275             


Arg Val  Ile Leu Ala Asp Ala  Asn Leu Asp Lys Val  Leu Ser Ala 
    1280                 1285                 1290             


Tyr Asn  Lys His Arg Asp Lys  Pro Ile Arg Glu Gln  Ala Glu Asn 
    1295                 1300                 1305             


Ile Ile  His Leu Phe Thr Leu  Thr Asn Leu Gly Ala  Pro Ala Ala 
    1310                 1315                 1320             


Phe Lys  Tyr Phe Asp Thr Thr  Ile Asp Arg Lys Arg  Tyr Thr Ser 
    1325                 1330                 1335             


Thr Lys  Glu Val Leu Asp Ala  Thr Leu Ile His Gln  Ser Ile Thr 
    1340                 1345                 1350             


Gly Leu  Tyr Glu Thr Arg Ile  Asp Leu Ser Gln Leu  Gly Gly Asp 
    1355                 1360                 1365             


<210>  12
<211>  1368
<212>  PRT
<213>  Streptococcus pyogenes


<220>
<221>  MISC_FEATURE
<223>  S. Pyogenes Cas9 D10A

<400>  12

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 Asp His 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 Asn 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 
    1010                 1015                 1020             


Lys Ser  Glu Gln Glu Ile Gly  Lys Ala Thr Ala Lys  Tyr Phe Phe 
    1025                 1030                 1035             


Tyr Ser  Asn Ile Met Asn Phe  Phe Lys Thr Glu Ile  Thr Leu Ala 
    1040                 1045                 1050             


Asn Gly  Glu Ile Arg Lys Arg  Pro Leu Ile Glu Thr  Asn Gly Glu 
    1055                 1060                 1065             


Thr Gly  Glu Ile Val Trp Asp  Lys Gly Arg Asp Phe  Ala Thr Val 
    1070                 1075                 1080             


Arg Lys  Val Leu Ser Met Pro  Gln Val Asn Ile Val  Lys Lys Thr 
    1085                 1090                 1095             


Glu Val  Gln Thr Gly Gly Phe  Ser Lys Glu Ser Ile  Leu Pro Lys 
    1100                 1105                 1110             


Arg Asn  Ser Asp Lys Leu Ile  Ala Arg Lys Lys Asp  Trp Asp Pro 
    1115                 1120                 1125             


Lys Lys  Tyr Gly Gly Phe Asp  Ser Pro Thr Val Ala  Tyr Ser Val 
    1130                 1135                 1140             


Leu Val  Val Ala Lys Val Glu  Lys Gly Lys Ser Lys  Lys Leu Lys 
    1145                 1150                 1155             


Ser Val  Lys Glu Leu Leu Gly  Ile Thr Ile Met Glu  Arg Ser Ser 
    1160                 1165                 1170             


Phe Glu  Lys Asn Pro Ile Asp  Phe Leu Glu Ala Lys  Gly Tyr Lys 
    1175                 1180                 1185             


Glu Val  Lys Lys Asp Leu Ile  Ile Lys Leu Pro Lys  Tyr Ser Leu 
    1190                 1195                 1200             


Phe Glu  Leu Glu Asn Gly Arg  Lys Arg Met Leu Ala  Ser Ala Gly 
    1205                 1210                 1215             


Glu Leu  Gln Lys Gly Asn Glu  Leu Ala Leu Pro Ser  Lys Tyr Val 
    1220                 1225                 1230             


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 
    1250                 1255                 1260             


His Tyr  Leu Asp Glu Ile Ile  Glu Gln Ile Ser Glu  Phe Ser Lys 
    1265                 1270                 1275             


Arg Val  Ile Leu Ala Asp Ala  Asn Leu Asp Lys Val  Leu Ser Ala 
    1280                 1285                 1290             


Tyr Asn  Lys His Arg Asp Lys  Pro Ile Arg Glu Gln  Ala Glu Asn 
    1295                 1300                 1305             


Ile Ile  His Leu Phe Thr Leu  Thr Asn Leu Gly Ala  Pro Ala Ala 
    1310                 1315                 1320             


Phe Lys  Tyr Phe Asp Thr Thr  Ile Asp Arg Lys Arg  Tyr Thr Ser 
    1325                 1330                 1335             


Thr Lys  Glu Val Leu Asp Ala  Thr Leu Ile His Gln  Ser Ile Thr 
    1340                 1345                 1350             


Gly Leu  Tyr Glu Thr Arg Ile  Asp Leu Ser Gln Leu  Gly Gly Asp 
    1355                 1360                 1365             


<210>  13
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  PDCD1 CRISPR guide RNA target sequence 1

<400>  13
tgacgttacc tcgtgcggcc                                                   20


<210>  14
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  PDCD1 CRISPR guide RNA target sequence 2

<400>  14
cacgaagctc tccgatgtgt                                                   20


<210>  15
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  PDCD1 CRISPR guide RNA target sequence 3

<400>  15
gcgtgacttc cacatgagcg                                                   20


<210>  16
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  PDCD1 CRISPR guide RNA target sequence 4

<400>  16
ttggaactgg ccggctggcc                                                   20


<210>  17
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  PDCD1 CRISPR guide RNA target sequence 5

<400>  17
gtggcatact ccgtctgctc                                                   20


<210>  18
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  PDCD1 CRISPR guide RNA target sequence 6

<400>  18
gatgaggtgc ccattccgct                                                   20


<210>  19
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  CD274 CRISPR guide RNA target sequence 1

<400>  19
taccgctgca tgatcagcta                                                   20


<210>  20
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  CD274 CRISPR guide RNA target sequence 2

<400>  20
agctactatg ctgaaccttc                                                   20


<210>  21
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  CD274 CRISPR guide RNA target sequence 3

<400>  21
ggatgaccaa ttcagctgta                                                   20


<210>  22
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  CD274 CRISPR guide RNA target sequence 4

<400>  22
accccaaggc cgaagtcatc                                                   20


<210>  23
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  CD274 CRISPR guide RNA target sequence 5

<400>  23
tctttatatt catgacctac                                                   20


<210>  24
<211>  20
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  CD274 CRISPR guide RNA target sequence 6

<400>  24
accgttcagc aaatgccagt                                                   20


