                         SEQUENCE LISTING

<110>  New England Biolabs, Inc.
       Gardner, Andrew
 
<120>  Exonuclease-Deficient DNA Polymerase Variants and Uses Thereof

<130>  NEB-343-PCT

<150>  61/484,731
<151>  2011-05-11

<160>  12    

<170>  PatentIn version 3.5

<210>  1
<211>  775
<212>  PRT
<213>  Thermococcus sp.


<220>
<221>  MISC_FEATURE
<222>  (1)..(755)
<223>  9 degrees N strain

<400>  1

Met Ile Leu Asp Thr Asp Tyr Ile Thr Glu Asn Gly Lys Pro Val Ile 
1               5                   10                  15      


Arg Val Phe Lys Lys Glu Asn Gly Glu Phe Lys Ile Glu Tyr Asp Arg 
            20                  25                  30          


Thr Phe Glu Pro Tyr Phe Tyr Ala Leu Leu Lys Asp Asp Ser Ala Ile 
        35                  40                  45              


Glu Asp Val Lys Lys Val Thr Ala Lys Arg His Gly Thr Val Val Lys 
    50                  55                  60                  


Val Lys Arg Ala Glu Lys Val Gln Lys Lys Phe Leu Gly Arg Pro Ile 
65                  70                  75                  80  


Glu Val Trp Lys Leu Tyr Phe Asn His Pro Gln Asp Val Pro Ala Ile 
                85                  90                  95      


Arg Asp Arg Ile Arg Ala His Pro Ala Val Val Asp Ile Tyr Glu Tyr 
            100                 105                 110         


Asp Ile Pro Phe Ala Lys Arg Tyr Leu Ile Asp Lys Gly Leu Ile Pro 
        115                 120                 125             


Met Glu Gly Asp Glu Glu Leu Thr Met Leu Ala Phe Asp Ile Glu Thr 
    130                 135                 140                 


Leu Tyr His Glu Gly Glu Glu Phe Gly Thr Gly Pro Ile Leu Met Ile 
145                 150                 155                 160 


Ser Tyr Ala Asp Gly Ser Glu Ala Arg Val Ile Thr Trp Lys Lys Ile 
                165                 170                 175     


Asp Leu Pro Tyr Val Asp Val Val Ser Thr Glu Lys Glu Met Ile Lys 
            180                 185                 190         


Arg Phe Leu Arg Val Val Arg Glu Lys Asp Pro Asp Val Leu Ile Thr 
        195                 200                 205             


Tyr Asn Gly Asp Asn Phe Asp Phe Ala Tyr Leu Lys Lys Arg Cys Glu 
    210                 215                 220                 


Glu Leu Gly Ile Lys Phe Thr Leu Gly Arg Asp Gly Ser Glu Pro Lys 
225                 230                 235                 240 


Ile Gln Arg Met Gly Asp Arg Phe Ala Val Glu Val Lys Gly Arg Ile 
                245                 250                 255     


His Phe Asp Leu Tyr Pro Val Ile Arg Arg Thr Ile Asn Leu Pro Thr 
            260                 265                 270         


Tyr Thr Leu Glu Ala Val Tyr Glu Ala Val Phe Gly Lys Pro Lys Glu 
        275                 280                 285             


Lys Val Tyr Ala Glu Glu Ile Ala Gln Ala Trp Glu Ser Gly Glu Gly 
    290                 295                 300                 


Leu Glu Arg Val Ala Arg Tyr Ser Met Glu Asp Ala Lys Val Thr Tyr 
305                 310                 315                 320 


Glu Leu Gly Arg Glu Phe Phe Pro Met Glu Ala Gln Leu Ser Arg Leu 
                325                 330                 335     


Ile Gly Gln Ser Leu Trp Asp Val Ser Arg Ser Ser Thr Gly Asn Leu 
            340                 345                 350         


Val Glu Trp Phe Leu Leu Arg Lys Ala Tyr Lys Arg Asn Glu Leu Ala 
        355                 360                 365             


Pro Asn Lys Pro Asp Glu Arg Glu Leu Ala Arg Arg Arg Gly Gly Tyr 
    370                 375                 380                 


Ala Gly Gly Tyr Val Lys Glu Pro Glu Arg Gly Leu Trp Asp Asn Ile 
385                 390                 395                 400 


Val Tyr Leu Asp Phe Arg Ser Leu Tyr Pro Ser Ile Ile Ile Thr His 
                405                 410                 415     


Asn Val Ser Pro Asp Thr Leu Asn Arg Glu Gly Cys Lys Glu Tyr Asp 
            420                 425                 430         


Val Ala Pro Glu Val Gly His Lys Phe Cys Lys Asp Phe Pro Gly Phe 
        435                 440                 445             


Ile Pro Ser Leu Leu Gly Asp Leu Leu Glu Glu Arg Gln Lys Ile Lys 
    450                 455                 460                 


Arg Lys Met Lys Ala Thr Val Asp Pro Leu Glu Lys Lys Leu Leu Asp 
465                 470                 475                 480 


Tyr Arg Gln Arg Ala Ile Lys Ile Leu Ala Asn Ser Phe Tyr Gly Tyr 
                485                 490                 495     


Tyr Gly Tyr Ala Lys Ala Arg Trp Tyr Cys Lys Glu Cys Ala Glu Ser 
            500                 505                 510         


Val Thr Ala Trp Gly Arg Glu Tyr Ile Glu Met Val Ile Arg Glu Leu 
        515                 520                 525             


Glu Glu Lys Phe Gly Phe Lys Val Leu Tyr Ala Asp Thr Asp Gly Leu 
    530                 535                 540                 


His Ala Thr Ile Pro Gly Ala Asp Ala Glu Thr Val Lys Lys Lys Ala 
545                 550                 555                 560 


Lys Glu Phe Leu Lys Tyr Ile Asn Pro Lys Leu Pro Gly Leu Leu Glu 
                565                 570                 575     


Leu Glu Tyr Glu Gly Phe Tyr Val Arg Gly Phe Phe Val Thr Lys Lys 
            580                 585                 590         


Lys Tyr Ala Val Ile Asp Glu Glu Gly Lys Ile Thr Thr Arg Gly Leu 
        595                 600                 605             


Glu Ile Val Arg Arg Asp Trp Ser Glu Ile Ala Lys Glu Thr Gln Ala 
    610                 615                 620                 


Arg Val Leu Glu Ala Ile Leu Lys His Gly Asp Val Glu Glu Ala Val 
625                 630                 635                 640 


Arg Ile Val Lys Glu Val Thr Glu Lys Leu Ser Lys Tyr Glu Val Pro 
                645                 650                 655     


Pro Glu Lys Leu Val Ile His Glu Gln Ile Thr Arg Asp Leu Arg Asp 
            660                 665                 670         


Tyr Lys Ala Thr Gly Pro His Val Ala Val Ala Lys Arg Leu Ala Ala 
        675                 680                 685             


Arg Gly Val Lys Ile Arg Pro Gly Thr Val Ile Ser Tyr Ile Val Leu 
    690                 695                 700                 


Lys Gly Ser Gly Arg Ile Gly Asp Arg Ala Ile Pro Ala Asp Glu Phe 
705                 710                 715                 720 


Asp Pro Thr Lys His Arg Tyr Asp Ala Glu Tyr Tyr Ile Glu Asn Gln 
                725                 730                 735     


Val Leu Pro Ala Val Glu Arg Ile Leu Lys Ala Phe Gly Tyr Arg Lys 
            740                 745                 750         


Glu Asp Leu Arg Tyr Gln Lys Thr Lys Gln Val Gly Leu Gly Ala Trp 
        755                 760                 765             


Leu Lys Val Lys Gly Lys Lys 
    770                 775 


<210>  2
<211>  774
<212>  PRT
<213>  Thermococcus litoralis

<400>  2

Met Ile Leu Asp Thr Asp Tyr Ile Thr Lys Asp Gly Lys Pro Ile Ile 
1               5                   10                  15      


Arg Ile Phe Lys Lys Glu Asn Gly Glu Phe Lys Ile Glu Leu Asp Pro 
            20                  25                  30          


His Phe Gln Pro Tyr Ile Tyr Ala Leu Leu Lys Asp Asp Ser Ala Ile 
        35                  40                  45              


Glu Glu Ile Lys Ala Ile Lys Gly Glu Arg His Gly Lys Thr Val Arg 
    50                  55                  60                  


Val Leu Asp Ala Val Lys Val Arg Lys Lys Phe Leu Gly Arg Glu Val 
65                  70                  75                  80  


Glu Val Trp Lys Leu Ile Phe Glu His Pro Gln Asp Val Pro Ala Met 
                85                  90                  95      


Arg Gly Lys Ile Arg Glu His Pro Ala Val Val Asp Ile Tyr Glu Tyr 
            100                 105                 110         


Asp Ile Pro Phe Ala Lys Arg Tyr Leu Ile Asp Lys Gly Leu Ile Pro 
        115                 120                 125             


Met Glu Gly Asp Glu Glu Leu Lys Leu Leu Ala Phe Asp Ile Glu Thr 
    130                 135                 140                 


Phe Tyr His Glu Gly Asp Glu Phe Gly Lys Gly Glu Ile Ile Met Ile 
145                 150                 155                 160 


Ser Tyr Ala Asp Glu Glu Glu Ala Arg Val Ile Thr Trp Lys Asn Ile 
                165                 170                 175     


Asp Leu Pro Tyr Val Asp Val Val Ser Asn Glu Arg Glu Met Ile Lys 
            180                 185                 190         


Arg Phe Val Gln Val Val Lys Glu Lys Asp Pro Asp Val Ile Ile Thr 
        195                 200                 205             


Tyr Asn Gly Asp Asn Phe Asp Leu Pro Tyr Leu Ile Lys Arg Ala Glu 
    210                 215                 220                 


Lys Leu Gly Val Arg Leu Val Leu Gly Arg Asp Lys Glu His Pro Glu 
225                 230                 235                 240 


Pro Lys Ile Gln Arg Met Gly Asp Ser Phe Ala Val Glu Ile Lys Gly 
                245                 250                 255     


Arg Ile His Phe Asp Leu Phe Pro Val Val Arg Arg Thr Ile Asn Leu 
            260                 265                 270         


Pro Thr Tyr Thr Leu Glu Ala Val Tyr Glu Ala Val Leu Gly Lys Thr 
        275                 280                 285             


Lys Ser Lys Leu Gly Ala Glu Glu Ile Ala Ala Ile Trp Glu Thr Glu 
    290                 295                 300                 


Glu Ser Met Lys Lys Leu Ala Gln Tyr Ser Met Glu Asp Ala Arg Ala 
305                 310                 315                 320 


Thr Tyr Glu Leu Gly Lys Glu Phe Phe Pro Met Glu Ala Glu Leu Ala 
                325                 330                 335     


Lys Leu Ile Gly Gln Ser Val Trp Asp Val Ser Arg Ser Ser Thr Gly 
            340                 345                 350         


Asn Leu Val Glu Trp Tyr Leu Leu Arg Val Ala Tyr Ala Arg Asn Glu 
        355                 360                 365             


Leu Ala Pro Asn Lys Pro Asp Glu Glu Glu Tyr Lys Arg Arg Leu Arg 
    370                 375                 380                 


Thr Thr Tyr Leu Gly Gly Tyr Val Lys Glu Pro Glu Lys Gly Leu Trp 
385                 390                 395                 400 


Glu Asn Ile Ile Tyr Leu Asp Phe Arg Ser Leu Tyr Pro Ser Ile Ile 
                405                 410                 415     


Val Thr His Asn Val Ser Pro Asp Thr Leu Glu Lys Glu Gly Cys Lys 
            420                 425                 430         


Asn Tyr Asp Val Ala Pro Ile Val Gly Tyr Arg Phe Cys Lys Asp Phe 
        435                 440                 445             


Pro Gly Phe Ile Pro Ser Ile Leu Gly Asp Leu Ile Ala Met Arg Gln 
    450                 455                 460                 


Asp Ile Lys Lys Lys Met Lys Ser Thr Ile Asp Pro Ile Glu Lys Lys 
465                 470                 475                 480 


Met Leu Asp Tyr Arg Gln Arg Ala Ile Lys Leu Leu Ala Asn Ser Tyr 
                485                 490                 495     


Tyr Gly Tyr Met Gly Tyr Pro Lys Ala Arg Trp Tyr Ser Lys Glu Cys 
            500                 505                 510         


Ala Glu Ser Val Thr Ala Trp Gly Arg His Tyr Ile Glu Met Thr Ile 
        515                 520                 525             


Arg Glu Ile Glu Glu Lys Phe Gly Phe Lys Val Leu Tyr Ala Asp Thr 
    530                 535                 540                 


Asp Gly Phe Tyr Ala Thr Ile Pro Gly Glu Lys Pro Glu Leu Ile Lys 
545                 550                 555                 560 


Lys Lys Ala Lys Glu Phe Leu Asn Tyr Ile Asn Ser Lys Leu Pro Gly 
                565                 570                 575     


Leu Leu Glu Leu Glu Tyr Glu Gly Phe Tyr Leu Arg Gly Phe Phe Val 
            580                 585                 590         


Thr Lys Lys Arg Tyr Ala Val Ile Asp Glu Glu Gly Arg Ile Thr Thr 
        595                 600                 605             


Arg Gly Leu Glu Val Val Arg Arg Asp Trp Ser Glu Ile Ala Lys Glu 
    610                 615                 620                 


Thr Gln Ala Lys Val Leu Glu Ala Ile Leu Lys Glu Gly Ser Val Glu 
625                 630                 635                 640 


Lys Ala Val Glu Val Val Arg Asp Val Val Glu Lys Ile Ala Lys Tyr 
                645                 650                 655     


Arg Val Pro Leu Glu Lys Leu Val Ile His Glu Gln Ile Thr Arg Asp 
            660                 665                 670         


Leu Lys Asp Tyr Lys Ala Ile Gly Pro His Val Ala Ile Ala Lys Arg 
        675                 680                 685             


Leu Ala Ala Arg Gly Ile Lys Val Lys Pro Gly Thr Ile Ile Ser Tyr 
    690                 695                 700                 


Ile Val Leu Lys Gly Ser Gly Lys Ile Ser Asp Arg Val Ile Leu Leu 
705                 710                 715                 720 


Thr Glu Tyr Asp Pro Arg Lys His Lys Tyr Asp Pro Asp Tyr Tyr Ile 
                725                 730                 735     


Glu Asn Gln Val Leu Pro Ala Val Leu Arg Ile Leu Glu Ala Phe Gly 
            740                 745                 750         


Tyr Arg Lys Glu Asp Leu Arg Tyr Gln Ser Ser Lys Gln Thr Gly Leu 
        755                 760                 765             


Asp Ala Trp Leu Lys Arg 
    770                 


<210>  3
<211>  7
<212>  PRT
<213>  Thermococcus sp.


<220>
<221>  MISC_FEATURE
<222>  (1)..(7)
<223>  Exo Motif I corresponding to residues 138-144 in SEQ ID NO:1

<400>  3

Leu Ala Phe Asp Ile Glu Thr 
1               5           


<210>  4
<211>  9
<212>  PRT
<213>  Thermococcus sp.


<220>
<221>  MISC_FEATURE
<222>  (1)..(9)
<223>  Exo Motif II corresponding to residues 207-215 in SEQ ID NO:1

<400>  4

Ile Thr Tyr Asn Gly Asp Asn Phe Asp 
1               5                   


<210>  5
<211>  6
<212>  PRT
<213>  Thermococcus sp.


<220>
<221>  MISC_FEATURE
<222>  (1)..(6)
<223>  Exo Motif III corresponding to residues 311-316 in SEQ ID NO:1

<400>  5

Tyr Ser Met Glu Asp Ala 
1               5       


<210>  6
<211>  11
<212>  PRT
<213>  Thermococcus sp.


<220>
<221>  MISC_FEATURE
<222>  (1)..(11)
<223>  Exo Motif IV corresponding to residues 269-279 in SEQ ID NO:1

<220>
<221>  MISC_FEATURE
<222>  (9)..(9)
<223>  Xaa can be any naturally occurring amino acid


<400>  6

Asn Leu Pro Thr Tyr Thr Leu Glu Xaa Val Tyr 
1               5                   10      


<210>  7
<211>  6
<212>  PRT
<213>  Thermococcus sp.


<220>
<221>  MISC_FEATURE
<222>  (1)..(7)
<223>  Exo Motif V corresponding to residues 241-246 in SEQ ID NO:1

<400>  7

Ile Gln Arg Met Gly Asp 
1               5       


<210>  8
<211>  7
<212>  PRT
<213>  Thermococcus sp.


<220>
<221>  MISC_FEATURE
<222>  (1)..(7)
<223>  Exo Motif VI corresponding to residues 286-292 in SEQ ID NO:1

<400>  8

Pro Lys Glu Lys Val Tyr Ala 
1               5           


<210>  9
<211>  24
<212>  DNA
<213>  artificial

<220>
<223>  primer

<400>  9
cgccagggtt ttcccagtca cgac                                              24


<210>  10
<211>  44
<212>  DNA
<213>  artificial

<220>
<223>  template

<400>  10
aaccggttac gtacgtacgt gtcgtgactg ggaaaaccct ggcg                        44


<210>  11
<211>  12
<212>  PRT
<213>  unknown

<220>
<223>  motif, which can exist in many organisms


<220>
<221>  MISC_FEATURE
<222>  (1)..(1)
<223>  Xaa can be any non-polar amino acid (e.g., glycine, alanine, 
       valine, leucine, isoleucine, proline, methionine, phenylalanine, 
       tryptophan)

<220>
<221>  misc_feature
<222>  (2)..(2)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (5)..(5)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  MISC_FEATURE
<222>  (6)..(6)
<223>  Xaa can be any polar amino acid (e.g., serine, threonine, 
       asparagine, glutamine, cysteine, tyrosine, aspartic acid, 
       glutamic acid, lysine, arginine, histidine)

<220>
<221>  misc_feature
<222>  (7)..(7)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  MISC_FEATURE
<222>  (10)..(11)
<223>  Xaa can be any non-polar amino acid (e.g., glycine, alanine, 
       valine, leucine, isoleucine, proline, methionine, phenylalanine, 
       tryptophan)

<400>  11

Xaa Xaa Tyr Asn Xaa Xaa Xaa Phe Asp Xaa Xaa Tyr 
1               5                   10          


<210>  12
<211>  6
<212>  PRT
<213>  Thermococcus litoralis


<220>
<221>  MISC_FEATURE
<222>  (1)..(6)
<223>  corresponds to amino acid residues 287-292 in SEQ ID NO:2

<400>  12

Thr Lys Ser Lys Leu Gly 
1               5       


