                         SEQUENCE LISTING

<110>  New England Biolabs, Inc.
       Zhu, Zhenyu
       Guan, Shengxi
       Blanchard, Aine
 
<120>  Engineering a Novel Methylation-Specific Restriction Endonuclease

<130>  NEB-306-2-PCT

<150>  US 61/158,466
<151>  2009-03-09

<160>  9     

<170>  PatentIn version 3.5

<210>  1
<211>  1386
<212>  DNA
<213>  Bacillus stearothermophilus

<400>  1
atggagagtg aagctatgaa agtaatgaat acaggaaaca tgttagaaaa tcgtgatgaa       60

gctattcgtc aaatcttttt aaatatttta gaacaagatg aggatttttg ggattttcga      120

aatgaaaata ccaaggaatt ttcacacggg tatcattcat atccggccat gatgattcct      180

caagtagcaa gaaatttaat gcgtatgata ttgagtaatc aaccgaatat aaaatctgtt      240

ttcgaccctt ttatgggttc cggaaccact ttggttgaag gggttttaca tggtctagat      300

tcttttggta cggatttaaa tcctttggct agattattgg ggaaagttaa gactactcca      360

ttaaacccta cttatttatc ggaagttgct gaacaattta ttttttcttt aagtaatgat      420

gagttaagct ataaaaatgg tgaccttgtt ttagagaaac ctgaatttaa aaatattgat      480

tattggttta agccatatgt tattgattat ctacagatta tcaaaaataa catcaaaaaa      540

atacaacatg acgatatcaa gttattcttt tgggctgtat ttagtgaagt tgtgagatat      600

gtttctaaca ctagaaattc ggaatttaaa ttatatcgca tggatgaaga aaaattagcg      660

gattggaatc ctaatgtatt cgatgtattc attaagtttt taaatcgtaa tattgaattg      720

aaccagcaat tttatgaatt atataatgaa caaaacccta caaaccaacc agcggtaaaa      780

atattcgatt taaacgccat gaatctcgaa ggaattgatg ataacagttt cgatttactt      840

ataacatctc cgccttacgg tgatagtagg actacggttg cctatgggca attttcaagg      900

ttatctttac agtggttaga ttttaatgaa atcggggaag atgaatcttt agtcaaagac      960

ataaataaaa ttgatgcatt acttcttggt ggaaaagttg ataaagaatt aaaaaatagc     1020

ttagcttctg aaacgttaga aagaactata gaatcaattg ctatagaaga tgaaaaacgt     1080

gctaaagaag ttctgcaatt ctatattgat ttggataagg cacttaaaga aatagcaaga     1140

gttatgaaac ctaactcata tcaatgctgg gtagttggta atcgaacagt aaaaaaagta     1200

aagataccaa ctcatcaaat aataattgag ttattccaaa aatacggagt gagacacgtc     1260

ttaaccttcg aaagaaatat accaaacaaa aaaatgccta aagagaattc gccgacgaat     1320

aaggtcggtg agaaagttac tacaatgaac ggtgaagtga tttttattct aagaaaagaa     1380

ggataa                                                                1386


<210>  2
<211>  461
<212>  PRT
<213>  Bacillus stearothermophilus

<400>  2

Met Glu Ser Glu Ala Met Lys Val Met Asn Thr Gly Asn Met Leu Glu 
1               5                   10                  15      


Asn Arg Asp Glu Ala Ile Arg Gln Ile Phe Leu Asn Ile Leu Glu Gln 
            20                  25                  30          


Asp Glu Asp Phe Trp Asp Phe Arg Asn Glu Asn Thr Lys Glu Phe Ser 
        35                  40                  45              


His Gly Tyr His Ser Tyr Pro Ala Met Met Ile Pro Gln Val Ala Arg 
    50                  55                  60                  


Asn Leu Met Arg Met Ile Leu Ser Asn Gln Pro Asn Ile Lys Ser Val 
65                  70                  75                  80  


Phe Asp Pro Phe Met Gly Ser Gly Thr Thr Leu Val Glu Gly Val Leu 
                85                  90                  95      


His Gly Leu Asp Ser Phe Gly Thr Asp Leu Asn Pro Leu Ala Arg Leu 
            100                 105                 110         


Leu Gly Lys Val Lys Thr Thr Pro Leu Asn Pro Thr Tyr Leu Ser Glu 
        115                 120                 125             


Val Ala Glu Gln Phe Ile Phe Ser Leu Ser Asn Asp Glu Leu Ser Tyr 
    130                 135                 140                 


Lys Asn Gly Asp Leu Val Leu Glu Lys Pro Glu Phe Lys Asn Ile Asp 
145                 150                 155                 160 


Tyr Trp Phe Lys Pro Tyr Val Ile Asp Tyr Leu Gln Ile Ile Lys Asn 
                165                 170                 175     


Asn Ile Lys Lys Ile Gln His Asp Asp Ile Lys Leu Phe Phe Trp Ala 
            180                 185                 190         


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


Phe Lys Leu Tyr Arg Met Asp Glu Glu Lys Leu Ala Asp Trp Asn Pro 
    210                 215                 220                 


Asn Val Phe Asp Val Phe Ile Lys Phe Leu Asn Arg Asn Ile Glu Leu 
225                 230                 235                 240 


Asn Gln Gln Phe Tyr Glu Leu Tyr Asn Glu Gln Asn Pro Thr Asn Gln 
                245                 250                 255     


Pro Ala Val Lys Ile Phe Asp Leu Asn Ala Met Asn Leu Glu Gly Ile 
            260                 265                 270         


Asp Asp Asn Ser Phe Asp Leu Leu Ile Thr Ser Pro Pro Tyr Gly Asp 
        275                 280                 285             


Ser Arg Thr Thr Val Ala Tyr Gly Gln Phe Ser Arg Leu Ser Leu Gln 
    290                 295                 300                 


Trp Leu Asp Phe Asn Glu Ile Gly Glu Asp Glu Ser Leu Val Lys Asp 
305                 310                 315                 320 


Ile Asn Lys Ile Asp Ala Leu Leu Leu Gly Gly Lys Val Asp Lys Glu 
                325                 330                 335     


Leu Lys Asn Ser Leu Ala Ser Glu Thr Leu Glu Arg Thr Ile Glu Ser 
            340                 345                 350         


Ile Ala Ile Glu Asp Glu Lys Arg Ala Lys Glu Val Leu Gln Phe Tyr 
        355                 360                 365             


Ile Asp Leu Asp Lys Ala Leu Lys Glu Ile Ala Arg Val Met Lys Pro 
    370                 375                 380                 


Asn Ser Tyr Gln Cys Trp Val Val Gly Asn Arg Thr Val Lys Lys Val 
385                 390                 395                 400 


Lys Ile Pro Thr His Gln Ile Ile Ile Glu Leu Phe Gln Lys Tyr Gly 
                405                 410                 415     


Val Arg His Val Leu Thr Phe Glu Arg Asn Ile Pro Asn Lys Lys Met 
            420                 425                 430         


Pro Lys Glu Asn Ser Pro Thr Asn Lys Val Gly Glu Lys Val Thr Thr 
        435                 440                 445             


Met Asn Gly Glu Val Ile Phe Ile Leu Arg Lys Glu Gly 
    450                 455                 460     


<210>  3
<211>  711
<212>  DNA
<213>  Bacillus stearothermophilus

<400>  3
atgatggata ttaaaacatt catcaaaaaa ttggaagaaa tcaaagcaaa aggttacatt       60

cgtaccttga gacgtggaga tacaggagta ggtcatacgt tagaacagga acttggtttg      120

actgaaaaca acatctcttt accagattta ggtgtagcgg aacttaaagc ggctagacgc      180

aacacttctt caatgcttac acttttcact aaagaacctc tttctgataa aggtcgtaaa      240

cgtgaccgct atttattaga aacttttgct tatgatagcg ataaagaaga ccgtatcaaa      300

gaattgtaca ctacaataag cgctttggat tacaatgcac aaggttttaa attagaggtt      360

acaaataaag aaattcgtct cattcataaa gatataccat tagatgttta ctggacagca      420

gaattattac aaaaaacatt cgaagataaa cttccggcat tagtttatgt ttatgccgac      480

catataggag aagatgcaga cgaacatttc cattacacgg aagcacgttt actgaaagga      540

tttgatttta aaggttttat gaaagctgta caagatggtt atattaaagt cgatttgcgt      600

atgcacatga agaataacgg ccgacctcga aaccatggta cagctttcag aatcttacgg      660

agtcatctcc cgatttgttt taaagaacag caaattttag taaaaccata g               711


<210>  4
<211>  236
<212>  PRT
<213>  Bacillus stearothermophilus

<400>  4

Met Met Asp Ile Lys Thr Phe Ile Lys Lys Leu Glu Glu Ile Lys Ala 
1               5                   10                  15      


Lys Gly Tyr Ile Arg Thr Leu Arg Arg Gly Asp Thr Gly Val Gly His 
            20                  25                  30          


Thr Leu Glu Gln Glu Leu Gly Leu Thr Glu Asn Asn Ile Ser Leu Pro 
        35                  40                  45              


Asp Leu Gly Val Ala Glu Leu Lys Ala Ala Arg Arg Asn Thr Ser Ser 
    50                  55                  60                  


Met Leu Thr Leu Phe Thr Lys Glu Pro Leu Ser Asp Lys Gly Arg Lys 
65                  70                  75                  80  


Arg Asp Arg Tyr Leu Leu Glu Thr Phe Ala Tyr Asp Ser Asp Lys Glu 
                85                  90                  95      


Asp Arg Ile Lys Glu Leu Tyr Thr Thr Ile Ser Ala Leu Asp Tyr Asn 
            100                 105                 110         


Ala Gln Gly Phe Lys Leu Glu Val Thr Asn Lys Glu Ile Arg Leu Ile 
        115                 120                 125             


His Lys Asp Ile Pro Leu Asp Val Tyr Trp Thr Ala Glu Leu Leu Gln 
    130                 135                 140                 


Lys Thr Phe Glu Asp Lys Leu Pro Ala Leu Val Tyr Val Tyr Ala Asp 
145                 150                 155                 160 


His Ile Gly Glu Asp Ala Asp Glu His Phe His Tyr Thr Glu Ala Arg 
                165                 170                 175     


Leu Leu Lys Gly Phe Asp Phe Lys Gly Phe Met Lys Ala Val Gln Asp 
            180                 185                 190         


Gly Tyr Ile Lys Val Asp Leu Arg Met His Met Lys Asn Asn Gly Arg 
        195                 200                 205             


Pro Arg Asn His Gly Thr Ala Phe Arg Ile Leu Arg Ser His Leu Pro 
    210                 215                 220                 


Ile Cys Phe Lys Glu Gln Gln Ile Leu Val Lys Pro 
225                 230                 235     


<210>  5
<211>  54
<212>  DNA
<213>  artificial

<220>
<223>  primer

<400>  5
ggtggtggat ccggaggtac ctggatggag agtgaagcta tgaaagtaat gaat             54


<210>  6
<211>  48
<212>  DNA
<213>  artificial

<220>
<223>  primer

<400>  6
ggtggtgcat gcgcctggtt atccttcttt tcttagaata aaaatcac                    48


<210>  7
<211>  54
<212>  DNA
<213>  artificial

<220>
<223>  primer

<400>  7
ggtggtctgc agggaggtaa ataaatggat aaagaattaa aaaattatat ggat             54


<210>  8
<211>  42
<212>  DNA
<213>  artificial

<220>
<223>  primer

<400>  8
ggtggtggta ccctatggtt ttactaaaat ttgctgttct tt                          42


<210>  9
<211>  24
<212>  PRT
<213>  Bacillus stearothermophilus


<220>
<221>  misc_feature
<222>  (4)..(5)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (22)..(22)
<223>  Xaa can be any naturally occurring amino acid

<400>  9

Met Met Asp Xaa Xaa Lys Thr Phe Ile Lys Lys Leu Glu Glu Ile Lys 
1               5                   10                  15      


Ala Lys Gly Tyr Ile Xaa Thr Leu 
            20                  


