
                                SEQUENCE LISTING

<110> Xie, Jiahua
      Kittur, Farooqahmed S.
      Hung, Chiu-Yueh
      

<120> METHODS FOR THE PRODUCTION OF
  ASIALO-ERYTHROPOIETIN IN PLANTS AND ITS PURIFICATION FROM
  PLANT TISSUES

<130> 59873-430837

<150> US 61/652,599       
<151> 2012-05-29  

<160> 13

<170> FastSEQ for Windows Version 4.0

<210> 1
<211> 636
<212> DNA
<213> Artificial Sequence

<220> 
<223> Artificial Sequence of human EPO fusion protein

<220> 
<221> misc_feature   
<222> (1)...(579)
<223> humam EPO domain coding region

<220> 
<221> misc_feature   
<222> (580)...(600)
<223> TEV protease cleavage domain coding region

<220> 
<221> misc_feature   
<222> (601)...(624)
<223> StrepII tag domain coding region

<220> 
<221> misc_feature   
<222> (625)...(636)
<223> KDEL domain coding region

<400> 1
atgggggtgc acgaatgtcc tgcctggctg tggcttctcc tgtccctgct gtcgctccct 60
ctgggcctcc cagtcctggg cgccccacca cgcctcatct gtgacagccg agtcctggag 120
aggtacctct tggaggccaa ggaggccgag aatatcacga cgggctgtgc tgaacactgc 180
agcttgaatg agaatatcac tgtcccagac accaaagtta atttctatgc ctggaagagg 240
atggaggtcg ggcagcaggc cgtagaagtc tggcagggcc tggccctgct gtcggaagct 300
gtcctgcggg gccaggccct gttggtcaac tcttcccagc cgtgggagcc cctgcagctg 360
catgtggata aagccgtcag tggccttcgc agcctcacca ctctgcttcg ggctctggga 420
gcccagaagg aagccatctc ccctccagat gcggcctcag ctgctccact ccgaacaatc 480
actgctgaca ctttccgcaa actcttccga gtctactcca atttcctccg gggaaagctg 540
aagctgtaca caggggaggc ctgcaggaca ggggacagag aaaacctgta ttttcagggc 600
tggagtcatc ctcaatttga gaagaaagat gaactc                           636

<210> 2
<211> 212
<212> PRT
<213> Artificial Sequence

<220> 
<223> Artificial Sequence of human EPO fusion protein

<220> 
<221> DOMAIN         
<222> (1)...(193)
<223> human EPO domain

<220> 
<221> DOMAIN         
<222> (194)...(200)
<223> TEV protease cleavage domain

<220> 
<221> DOMAIN         
<222> (201)...(208)
<223> StrepII tag domain

<220> 
<221> DOMAIN         
<222> (209)...(212)
<223> KDEL endoplasmic reticulum retention signal domain

<400> 2
Met Gly Val His Glu Cys Pro Ala Trp Leu Trp Leu Leu Leu Ser Leu
 1               5                  10                  15      
Leu Ser Leu Pro Leu Gly Leu Pro Val Leu Gly Ala Pro Pro Arg Leu
            20                  25                  30          
Ile Cys Asp Ser Arg Val Leu Glu Arg Tyr Leu Leu Glu Ala Lys Glu
        35                  40                  45              
Ala Glu Asn Ile Thr Thr Gly Cys Ala Glu His Cys Ser Leu Asn Glu
    50                  55                  60                  
Asn Ile Thr Val Pro Asp Thr Lys Val Asn Phe Tyr Ala Trp Lys Arg
65                  70                  75                  80  
Met Glu Val Gly Gln Gln Ala Val Glu Val Trp Gln Gly Leu Ala Leu
                85                  90                  95      
Leu Ser Glu Ala Val Leu Arg Gly Gln Ala Leu Leu Val Asn Ser Ser
            100                 105                 110         
Gln Pro Trp Glu Pro Leu Gln Leu His Val Asp Lys Ala Val Ser Gly
        115                 120                 125             
Leu Arg Ser Leu Thr Thr Leu Leu Arg Ala Leu Gly Ala Gln Lys Glu
    130                 135                 140                 
Ala Ile Ser Pro Pro Asp Ala Ala Ser Ala Ala Pro Leu Arg Thr Ile
145                 150                 155                 160 
Thr Ala Asp Thr Phe Arg Lys Leu Phe Arg Val Tyr Ser Asn Phe Leu
                165                 170                 175     
Arg Gly Lys Leu Lys Leu Tyr Thr Gly Glu Ala Cys Arg Thr Gly Asp
            180                 185                 190         
Arg Glu Asn Leu Tyr Phe Gln Gly Trp Ser His Pro Gln Phe Glu Lys
        195                 200                 205             
Lys Asp Glu Leu
    210         


<210> 3
<211> 579
<212> DNA
<213> Homo sapiens

<220> 
<221> misc_feature   
<222> (1)...(579)
<223> Coding region of human EPO of Accession No.
      NM_000799

<400> 3
atgggggtgc acgaatgtcc tgcctggctg tggcttctcc tgtccctgct gtcgctccct 60
ctgggcctcc cagtcctggg cgccccacca cgcctcatct gtgacagccg agtcctggag 120
aggtacctct tggaggccaa ggaggccgag aatatcacga cgggctgtgc tgaacactgc 180
agcttgaatg agaatatcac tgtcccagac accaaagtta atttctatgc ctggaagagg 240
atggaggtcg ggcagcaggc cgtagaagtc tggcagggcc tggccctgct gtcggaagct 300
gtcctgcggg gccaggccct gttggtcaac tcttcccagc cgtgggagcc cctgcagctg 360
catgtggata aagccgtcag tggccttcgc agcctcacca ctctgcttcg ggctctggga 420
gcccagaagg aagccatctc ccctccagat gcggcctcag ctgctccact ccgaacaatc 480
actgctgaca ctttccgcaa actcttccga gtctactcca atttcctccg gggaaagctg 540
aagctgtaca caggggaggc ctgcaggaca ggggacaga                        579

<210> 4
<211> 193
<212> PRT
<213> Homo sapiens

<220> 
<221> DOMAIN         
<222> (1)...(193)
<223> human EPO protein of Accession No. NP_000790

<400> 4
Met Gly Val His Glu Cys Pro Ala Trp Leu Trp Leu Leu Leu Ser Leu
 1               5                  10                  15      
Leu Ser Leu Pro Leu Gly Leu Pro Val Leu Gly Ala Pro Pro Arg Leu
            20                  25                  30          
Ile Cys Asp Ser Arg Val Leu Glu Arg Tyr Leu Leu Glu Ala Lys Glu
        35                  40                  45              
Ala Glu Asn Ile Thr Thr Gly Cys Ala Glu His Cys Ser Leu Asn Glu
    50                  55                  60                  
Asn Ile Thr Val Pro Asp Thr Lys Val Asn Phe Tyr Ala Trp Lys Arg
65                  70                  75                  80  
Met Glu Val Gly Gln Gln Ala Val Glu Val Trp Gln Gly Leu Ala Leu
                85                  90                  95      
Leu Ser Glu Ala Val Leu Arg Gly Gln Ala Leu Leu Val Asn Ser Ser
            100                 105                 110         
Gln Pro Trp Glu Pro Leu Gln Leu His Val Asp Lys Ala Val Ser Gly
        115                 120                 125             
Leu Arg Ser Leu Thr Thr Leu Leu Arg Ala Leu Gly Ala Gln Lys Glu
    130                 135                 140                 
Ala Ile Ser Pro Pro Asp Ala Ala Ser Ala Ala Pro Leu Arg Thr Ile
145                 150                 155                 160 
Thr Ala Asp Thr Phe Arg Lys Leu Phe Arg Val Tyr Ser Asn Phe Leu
                165                 170                 175     
Arg Gly Lys Leu Lys Leu Tyr Thr Gly Glu Ala Cys Arg Thr Gly Asp
            180                 185                 190         
Arg
    


<210> 5
<211> 7
<212> PRT
<213> Artificial Sequence

<220> 
<223> TEV protease cleavage domain

<400> 5
Glu Asn Leu Tyr Phe Gln Gly
 1               5          


<210> 6
<211> 8
<212> PRT
<213> Artificial Sequence

<220> 
<223> StepII tag domain

<400> 6
Trp Ser His Pro Gln Phe Glu Lys
 1               5              


<210> 7
<211> 4
<212> PRT
<213> Artificial Sequence

<220> 
<223> KDEL endoplasmic reticulum retention signal domain

<400> 7
Lys Asp Glu Leu
 1              


<210> 8
<211> 4
<212> PRT
<213> Artificial Sequence

<220> 
<223> HDEL endoplasmic reticulum retention signal domain

<400> 8
His Asp Glu Leu
 1              


<210> 9
<211> 784
<212> DNA
<213> Artificial Sequence

<220> 
<223> Double CaMV 35S promoter

<400> 9
gaaaatcttc gtcaacatgg tggagcacga cacgcttgtc tacctccaaa aatatcaaag 60
atacagtctc agaagaccaa agggaattga gacttttcaa caaagggtaa tatccggaaa 120
cctcctcgga ttccattgcc cagctatctg tcactttatt gtgaagatag tggaaaagga 180
aggtggctcc tacaaatgcc atcattgcga taaaggaaag gccatcgttg aagatgcctc 240
tgccgacagt ggtcccaaag atggaccccc acccacgagg agcatcgtgg aaaaagaaga 300
cgttccaacc acgtcttcaa agcaagtgga ttgatgtgat aacatggtgg agcacgacac 360
gcttgtctac ctccaaaaat atcaaagata cagtctcaga agaccaaagg gaattgagac 420
ttttcaacaa agggtaatat ccggaaacct cctcggattc cattgcccag ctatctgtca 480
ctttattgtg aagatagtgg aaaaggaagg tggctcctac aaatgccatc attgcgataa 540
aggaaaggcc atcgttgaag atgcctctgc cgacagtggt cccaaagatg gacccccacc 600
cacgaggagc atcgtggaaa aagaagacgt tccaaccacg tcttcaaagc aagtggattg 660
atgtgatatc tccactgacg taagggatga cgcacaatcc cactatcctt cgcaagaccc 720
ttcctctata taaggaagtt catttcattt ggagaggaca cgctgaaatc accagtctct 780
ctct                                                              784

<210> 10
<211> 1194
<212> DNA
<213> Homo sapiens

<220> 
<221> misc_feature   
<222> (1)...(1194)
<223> Human beta 1,4- galactosyltransferase 1 DNA
      sequence of beta 1,4- galactosyltransferase 1:
      coding region of Acession No. NM_001497

<400> 10
atgaggcttc gggagccgct cctgagcggc agcgccgcga tgccaggcgc gtccctacag 60
cgggcctgcc gcctgctcgt ggccgtctgc gctctgcacc ttggcgtcac cctcgtttac 120
tacctggctg gccgcgacct gagccgcctg ccccaactgg tcggagtctc cacaccgctg 180
cagggcggct cgaacagtgc cgccgccatc gggcagtcct ccggggagct ccggaccgga 240
ggggcccggc cgccgcctcc tctaggcgcc tcctcccagc cgcgcccggg tggcgactcc 300
agcccagtcg tggattctgg ccctggcccc gctagcaact tgacctcggt cccagtgccc 360
cacaccaccg cactgtcgct gcccgcctgc cctgaggagt ccccgctgct tgtgggcccc 420
atgctgattg agtttaacat gcctgtggac ctggagctcg tggcaaagca gaacccaaat 480
gtgaagatgg gcggccgcta tgcccccagg gactgcgtct ctcctcacaa ggtggccatc 540
atcattccat tccgcaaccg gcaggagcac ctcaagtact ggctatatta tttgcaccca 600
gtcctgcagc gccagcagct ggactatggc atctatgtta tcaaccaggc gggagacact 660
atattcaatc gtgctaagct cctcaatgtt ggctttcaag aagccttgaa ggactatgac 720
tacacctgct ttgtgtttag tgacgtggac ctcattccaa tgaatgacca taatgcgtac 780
aggtgttttt cacagccacg gcacatttcc gttgcaatgg ataagtttgg attcagccta 840
ccttatgttc agtattttgg aggtgtctct gctctaagta aacaacagtt tctaaccatc 900
aatggatttc ctaataatta ttggggctgg ggaggagaag atgatgacat ttttaacaga 960
ttagttttta gaggcatgtc tatatctcgc ccaaatgctg tggtcgggag gtgtcgcatg 1020
atccgccact caagagacaa gaaaaatgaa cccaatcctc agaggtttga ccgaattgca 1080
cacacaaagg agacaatgct ctctgatggt ttgaactcac tcacctacca ggtgctggat 1140
gtacagagat acccattgta tacccaaatc acagtggaca tcgggacacc gagc       1194

<210> 11
<211> 398
<212> PRT
<213> Homo sapiens

<220> 
<221> DOMAIN         
<222> (1)...(398)
<223> beta 1,4- galactosyltransferase 1 of Accession No.
      NP_001488

<400> 11
Met Arg Leu Arg Glu Pro Leu Leu Ser Gly Ser Ala Ala Met Pro Gly
 1               5                  10                  15      
Ala Ser Leu Gln Arg Ala Cys Arg Leu Leu Val Ala Val Cys Ala Leu
            20                  25                  30          
His Leu Gly Val Thr Leu Val Tyr Tyr Leu Ala Gly Arg Asp Leu Ser
        35                  40                  45              
Arg Leu Pro Gln Leu Val Gly Val Ser Thr Pro Leu Gln Gly Gly Ser
    50                  55                  60                  
Asn Ser Ala Ala Ala Ile Gly Gln Ser Ser Gly Glu Leu Arg Thr Gly
65                  70                  75                  80  
Gly Ala Arg Pro Pro Pro Pro Leu Gly Ala Ser Ser Gln Pro Arg Pro
                85                  90                  95      
Gly Gly Asp Ser Ser Pro Val Val Asp Ser Gly Pro Gly Pro Ala Ser
            100                 105                 110         
Asn Leu Thr Ser Val Pro Val Pro His Thr Thr Ala Leu Ser Leu Pro
        115                 120                 125             
Ala Cys Pro Glu Glu Ser Pro Leu Leu Val Gly Pro Met Leu Ile Glu
    130                 135                 140                 
Phe Asn Met Pro Val Asp Leu Glu Leu Val Ala Lys Gln Asn Pro Asn
145                 150                 155                 160 
Val Lys Met Gly Gly Arg Tyr Ala Pro Arg Asp Cys Val Ser Pro His
                165                 170                 175     
Lys Val Ala Ile Ile Ile Pro Phe Arg Asn Arg Gln Glu His Leu Lys
            180                 185                 190         
Tyr Trp Leu Tyr Tyr Leu His Pro Val Leu Gln Arg Gln Gln Leu Asp
        195                 200                 205             
Tyr Gly Ile Tyr Val Ile Asn Gln Ala Gly Asp Thr Ile Phe Asn Arg
    210                 215                 220                 
Ala Lys Leu Leu Asn Val Gly Phe Gln Glu Ala Leu Lys Asp Tyr Asp
225                 230                 235                 240 
Tyr Thr Cys Phe Val Phe Ser Asp Val Asp Leu Ile Pro Met Asn Asp
                245                 250                 255     
His Asn Ala Tyr Arg Cys Phe Ser Gln Pro Arg His Ile Ser Val Ala
            260                 265                 270         
Met Asp Lys Phe Gly Phe Ser Leu Pro Tyr Val Gln Tyr Phe Gly Gly
        275                 280                 285             
Val Ser Ala Leu Ser Lys Gln Gln Phe Leu Thr Ile Asn Gly Phe Pro
    290                 295                 300                 
Asn Asn Tyr Trp Gly Trp Gly Gly Glu Asp Asp Asp Ile Phe Asn Arg
305                 310                 315                 320 
Leu Val Phe Arg Gly Met Ser Ile Ser Arg Pro Asn Ala Val Val Gly
                325                 330                 335     
Arg Cys Arg Met Ile Arg His Ser Arg Asp Lys Lys Asn Glu Pro Asn
            340                 345                 350         
Pro Gln Arg Phe Asp Arg Ile Ala His Thr Lys Glu Thr Met Leu Ser
        355                 360                 365             
Asp Gly Leu Asn Ser Leu Thr Tyr Gln Val Leu Asp Val Gln Arg Tyr
    370                 375                 380                 
Pro Leu Tyr Thr Gln Ile Thr Val Asp Ile Gly Thr Pro Ser
385                 390                 395             


<210> 12
<211> 1291
<212> DNA
<213> Nicotiana tabacum

<220> 
<221> promoter       
<222> (1)...(1291)
<223> Tobacco glyceraldehyde-3-phosphate dehydrogenase
      gene (GapC) promoter

<400> 12
tctagaatgt tcgtgcgtca aatggataaa caaaaaaata gcataagtta gttttgttac 60
tcgagagtta tgtattataa ggtataggga aatgactcaa acataccact gaacttaacg 120
aaacgacgca tatatatact acttaactta acgaaaaagg ggtgagagtg gatgggtgct 180
ggtaaataat gaagggttta tataacgtca cgtgtcaaaa ttcgatagta gtagtttcgt 240
tagttgtaat agcatatatg gcccaaagtt ataatataga taatatgttt atgtccaact 300
attaacgagt gacatagaca gttcattttg tgaagttcaa tgacatattt gagccctttc 360
ccttttatta tctcctttta tttgttctaa taaaagaatg gcatttatta tgtacataga 420
caaataacta ttttctttgg aatataattt gtttatatat tttaaaatca tgtctcaatt 480
tagtttgttt tgtgcatatt tcaactattc aattttgtcc atatatttat taccttcccc 540
catttacaag cattgaaccg ctttgctcac caaaacttat gcacattgca aaaatatcat 600
gtaaaggttt tatgtatgct gtaattaagg tctgaactca tcgtgatttt atttttaggc 660
ttcattgacc actaccaaac tctttgatgc tacattttct aattatattg gagttcgatt 720
atatccgaat tcgcgttgcg tagggcccat tcgagggaaa acactcccta tcaaggattt 780
tttcataccc agagctcgaa ctcaagacat ctggttaagg gaagaacagt ctcatccact 840
gcaccatatc cttttgtggt caacaagtaa attttatgta gaaccaaaaa ctatactcga 900
attgataaaa taaatggtgt aaaatattgt tttctttctt acattttgga cagtaaatat 960
gtaggacaat aataattagc gtggggtctt aagaaaatta gcatagattt ccagaaattc 1020
caaatcaacc ggcagttcca ggtttgaaaa ctacaactca ttccgacggt tcaaacttca 1080
aaccatgctt gctgactcgg cttcttcttt ctttttcacc aagacagagc agtagtcacg 1140
tgacacccct cacgtgcctc ccccctttat atttcagact gcaaccctac actttcgcta 1200
cattcactac catattcttt tcactaagca attttctctc ctacttttct ttaaacccct 1260
tttttctccc ctaagccatg gcatctagat c                                1291

<210> 13
<211> 497
<212> DNA
<213> Nicotiana tabacum

<220> 
<221> terminator     
<222> (1)...(497)
<223> Tobacco GapC terminator

<400> 13
gagctcgtga aatggcctct ttagtttttg attgaatcat aggggtatta gttttctatg 60
gccgggagtg gtcttcttgc ttaattgtaa tggaataacc agagaggaac tactgtgtta 120
tctttgagga atgttgggct tttttcgttt gaattatcat gaatgaaatt ttactttttc 180
ccaatacaag tttgttttcg tttcttggtt tttgttatcc cttggtttat gtcttggttt 240
ggcttaaatg attgaagatt acactaccta tgtttctgct attcctgttg aagatcacat 300
ttgataataa tgcatcgaat gcattaaagt ttcttattgg ctctgtcaaa agtattgaag 360
gtggattttt ctaattggca agagaaagta ttaaagaggt gatttattag tacttatatt 420
tttctcagca tctctctttc agtgttggag cttcataaaa ttagcacttc agagtttcag 480
tcgggagctg aattcga                                                497
