                         SEQUENCE LISTING

<110>  VIRGINIA TECH INTELLECTUAL PROPERTIES, INC.
 
<120>  A MULTIVALENT BRUCELLA VACCINE FOR PROTECTION AGAINST 
       MYCOBACTERIAL INFECTIONS AND METHODS OF USING THE SAME

<130>  01640672TA

<160>  11    

<170>  PatentIn version 3.5

<210>  1
<211>  306
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Synthetic codon-optimized ESAT6

<400>  1
ggatccatga cggaacagca gtggaatttc gccggcatcg aagccgccgc ctccgccatc       60

cagggcaatg tgacgagcat ccatagcctc ctcgatgaag gcaagcagtc gctgaccaag      120

cttgccgcgg cctggggcgg ctcgggcagc gaagcgtatc agggcgtcca gcagaagtgg      180

gacgcgaccg ccacggaact taacaatgcg ctccagaacc tggcccgcac gatctcggaa      240

gcgggccagg cgatggcgtc cacggaaggc aatgtcacgg gcatgttcgc gagatcttaa      300

tctaga                                                                 306


<210>  2
<211>  876
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Synthetic codon-optimized Ag85B

<400>  2
ggatccttct cccgcccggg cctcccggtc gaatatctcc aggttccgtc cccgagcatg       60

ggccgcgaca tcaaggtcca gttccagtcc ggcggcaaca attcgccggc cgtctatctg      120

cttgacggcc ttcgcgcgca ggatgactat aatggctggg atatcaacac cccggccttc      180

gaatggtatt atcagagcgg cctgtccatc gtcatgccgg tgggcggcca gtcgagcttc      240

tattcggact ggtatagccc ggcctgcggc aaggcgggct gccagaccta taagtgggaa      300

accctcctga cgtcggaact cccgcagtgg ctgagcgcca atcgcgcggt gaagccgacg      360

ggctccgccg cgatcggcct ctcgatggcc ggctcctcgg cgatgatcct ggccgcgtat      420

catccgcagc agttcatcta tgccggctcc ctgtcggcgc ttctcgaccc gagccagggc      480

atgggcccgt ccctgatcgg ccttgccatg ggcgatgcgg gcggctataa ggccgcggat      540

atgtggggcc cgagctccga cccggcctgg gaacgcaacg atccgaccca gcagatcccg      600

aagcttgttg ccaacaatac ccgcctctgg gtctattgcg gcaacggcac gccgaatgaa      660

ctgggcggcg ccaatatccc ggcggaattc ctggaaaatt tcgtgcgctc gagcaacctt      720

aagttccagg atgcctataa cgccgcgggc ggccataacg cggttttcaa tttcccgccg      780

aacggcacgc actcgtggga atattggggc gcgcagctca atgccatgaa gggcgacctc      840

cagtcctccc tcggcgcggg cagatcttaa tctaga                                876


<210>  3
<211>  246
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Synthetic codon-optimized Rv2660c

<400>  3
ggatccgtta tcgcgggcgt cgatcaggcg cttgcggcca cgggccaggc gtcccagcgc       60

gcggcgggcg cgtcgggcgg cgttacggtc ggcgttggcg tcggcaccga acagcgcaac      120

ctgagcgtcg tggccccgtc gcagttcacg ttctcgtccc gctcgccgga tttcgtcgat      180

gaaacggcgg gccagtcgtg gtgcgcgatc ctcggcctta atcagttcca tagatcttga      240

tctaga                                                                 246


<210>  4
<211>  1147
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Synthetic trcD-Ag85B codon-optimized

<400>  4
gtcgaccaga aaaaagatca aaaaaatttg acaattaatc atccggctcg tataatgtgt       60

ggaattgtga gcggataaca atttcacaca ggaaacagcg ccgctgagaa aaagcgaagc      120

ggcactgctc tttaacaatt tatcagacaa tctgtgtggg cactcgaccg gaattatcga      180

ttaactttat tattaaaaat taaagaggta tatattaatg tatcgattaa ataaggagga      240

ataaaccatg catcatcatc atcatcatgg tggatccttc tcccgcccgg gcctcccggt      300

cgaatatctc caggttccgt ccccgagcat gggccgcgac atcaaggtcc agttccagtc      360

cggcggcaac aattcgccgg ccgtctatct gcttgacggc cttcgcgcgc aggatgacta      420

taatggctgg gatatcaaca ccccggcctt cgaatggtat tatcagagcg gcctgtccat      480

cgtcatgccg gtgggcggcc agtcgagctt ctattcggac tggtatagcc cggcctgcgg      540

caaggcgggc tgccagacct ataagtggga aaccctcctg acgtcggaac tcccgcagtg      600

gctgagcgcc aatcgcgcgg tgaagccgac gggctccgcc gcgatcggcc tctcgatggc      660

cggctcctcg gcgatgatcc tggccgcgta tcatccgcag cagttcatct atgccggctc      720

cctgtcggcg cttctcgacc cgagccaggg catgggcccg tccctgatcg gccttgccat      780

gggcgatgcg ggcggctata aggccgcgga tatgtggggc ccgagctccg acccggcctg      840

ggaacgcaac gatccgaccc agcagatccc gaagcttgtt gccaacaata cccgcctctg      900

ggtctattgc ggcaacggca cgccgaatga actgggcggc gccaatatcc cggcggaatt      960

cctggaaaat ttcgtgcgct cgagcaacct taagttccag gatgcctata acgccgcggg     1020

cggccataac gcggttttca atttcccgcc gaacggcacg cactcgtggg aatattgggg     1080

cgcgcagctc aatgccatga agggcgacct ccagtcctcc ctcggcgcgg gcagatctta     1140

atctaga                                                               1147


<210>  5
<211>  299
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Synthetic trcD-Ag85B matched amino acids

<400>  5

Met His His His His His His Gly Gly Ser Phe Ser Arg Pro Gly Leu 
1               5                   10                  15      


Pro Val Glu Tyr Leu Gln Val Pro Ser Pro Ser Met Gly Arg Asp Ile 
            20                  25                  30          


Lys Val Gln Phe Gln Ser Gly Gly Asn Asn Ser Pro Ala Val Tyr Leu 
        35                  40                  45              


Leu Asp Gly Leu Arg Ala Gln Asp Asp Tyr Asn Gly Trp Asp Ile Asn 
    50                  55                  60                  


Thr Pro Ala Phe Glu Trp Tyr Tyr Gln Ser Gly Leu Ser Ile Val Met 
65                  70                  75                  80  


Pro Val Gly Gly Gln Ser Ser Phe Tyr Ser Asp Trp Tyr Ser Pro Ala 
                85                  90                  95      


Cys Gly Lys Ala Gly Cys Gln Thr Tyr Lys Trp Glu Thr Leu Leu Thr 
            100                 105                 110         


Ser Glu Leu Pro Gln Trp Leu Ser Ala Asn Arg Ala Val Lys Pro Thr 
        115                 120                 125             


Gly Ser Ala Ala Ile Gly Leu Ser Met Ala Gly Ser Ser Ala Met Ile 
    130                 135                 140                 


Leu Ala Ala Tyr His Pro Gln Gln Phe Ile Tyr Ala Gly Ser Leu Ser 
145                 150                 155                 160 


Ala Leu Leu Asp Pro Ser Gln Gly Met Gly Pro Ser Leu Ile Gly Leu 
                165                 170                 175     


Ala Met Gly Asp Ala Gly Gly Tyr Lys Ala Ala Asp Met Trp Gly Pro 
            180                 185                 190         


Ser Ser Asp Pro Ala Trp Glu Arg Asn Asp Pro Thr Gln Gln Ile Pro 
        195                 200                 205             


Lys Leu Val Ala Asn Asn Thr Arg Leu Trp Val Tyr Cys Gly Asn Gly 
    210                 215                 220                 


Thr Pro Asn Glu Leu Gly Gly Ala Asn Ile Pro Ala Glu Phe Leu Glu 
225                 230                 235                 240 


Asn Phe Val Arg Ser Ser Asn Leu Lys Phe Gln Asp Ala Tyr Asn Ala 
                245                 250                 255     


Ala Gly Gly His Asn Ala Val Phe Asn Phe Pro Pro Asn Gly Thr His 
            260                 265                 270         


Ser Trp Glu Tyr Trp Gly Ala Gln Leu Asn Ala Met Lys Gly Asp Leu 
        275                 280                 285             


Gln Ser Ser Leu Gly Ala Gly Arg Ser Ser Arg 
    290                 295                 


<210>  6
<211>  808
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Synthetic trcD-Rv2660C::ESAT6 codon-optimized

<400>  6
gtcgaccaga aaaaagatca aaaaaatttg acaattaatc atccggctcg tataatgtgt       60

ggaattgtga gcggataaca atttcacaca ggaaacagcg ccgctgagaa aaagcgaagc      120

ggcactgctc tttaacaatt tatcagacaa tctgtgtggg cactcgaccg gaattatcga      180

ttaactttat tattaaaaat taaagaggta tatattaatg tatcgattaa ataaggagga      240

ataaaccatg catcatcatc atcatcatgg tggatccgtt atcgcgggcg tcgatcaggc      300

gcttgcggcc acgggccagg cgtcccagcg cgcggcgggc gcgtcgggcg gcgttacggt      360

cggcgttggc gtcggcaccg aacagcgcaa cctgagcgtc gtggccccgt cgcagttcac      420

gttctcgtcc cgctcgccgg atttcgtcga tgaaacggcg ggccagtcgt ggtgcgcgat      480

cctcggcctt aatcagttcc atagatccat gacggaacag cagtggaatt tcgccggcat      540

cgaagccgcc gcctccgcca tccagggcaa tgtgacgagc atccatagcc tcctcgatga      600

aggcaagcag tcgctgacca agcttgccgc ggcctggggc ggctcgggca gcgaagcgta      660

tcagggcgtc cagcagaagt gggacgcgac cgccacggaa cttaacaatg cgctccagaa      720

cctggcccgc acgatctcgg aagcgggcca ggcgatggcg tccacggaag gcaatgtcac      780

gggcatgttc gcgagatctt aatctaga                                         808


<210>  7
<211>  186
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Synthetic trcD-Rv2660c::ESAT6 matched amino acids

<400>  7

Met His His His His His His Gly Gly Ser Val Ile Ala Gly Val Asp 
1               5                   10                  15      


Gln Ala Leu Ala Ala Thr Gly Gln Ala Ser Gln Arg Ala Ala Gly Ala 
            20                  25                  30          


Ser Gly Gly Val Thr Val Gly Val Gly Val Gly Thr Glu Gln Arg Asn 
        35                  40                  45              


Leu Ser Val Val Ala Pro Ser Gln Phe Thr Phe Ser Ser Arg Ser Pro 
    50                  55                  60                  


Asp Phe Val Asp Glu Thr Ala Gly Gln Ser Trp Cys Ala Ile Leu Gly 
65                  70                  75                  80  


Leu Asn Gln Phe His Arg Ser Met Thr Glu Gln Gln Trp Asn Phe Ala 
                85                  90                  95      


Gly Ile Glu Ala Ala Ala Ser Ala Ile Gln Gly Asn Val Thr Ser Ile 
            100                 105                 110         


His Ser Leu Leu Asp Glu Gly Lys Gln Ser Leu Thr Lys Leu Ala Ala 
        115                 120                 125             


Ala Trp Gly Gly Ser Gly Ser Glu Ala Tyr Gln Gly Val Gln Gln Lys 
    130                 135                 140                 


Trp Asp Ala Thr Ala Thr Glu Leu Asn Asn Ala Leu Gln Asn Leu Ala 
145                 150                 155                 160 


Arg Thr Ile Ser Glu Ala Gly Gln Ala Met Ala Ser Thr Glu Gly Asn 
                165                 170                 175     


Val Thr Gly Met Phe Ala Arg Ser Ser Arg 
            180                 185     


<210>  8
<211>  297
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Synthetic protein product 6X His-tag-Ag85B

<400>  8

Met His His His His His His Gly Gly Ser Phe Ser Arg Pro Gly Leu 
1               5                   10                  15      


Pro Val Glu Tyr Leu Gln Val Pro Ser Pro Ser Met Gly Arg Asp Ile 
            20                  25                  30          


Lys Val Gln Phe Gln Ser Gly Gly Asn Asn Ser Pro Ala Val Tyr Leu 
        35                  40                  45              


Leu Asp Gly Leu Arg Ala Gln Asp Asp Tyr Asn Gly Trp Asp Ile Asn 
    50                  55                  60                  


Thr Pro Ala Phe Glu Trp Tyr Tyr Gln Ser Gly Leu Ser Ile Val Met 
65                  70                  75                  80  


Pro Val Gly Gly Gln Ser Ser Phe Tyr Ser Asp Trp Tyr Ser Pro Ala 
                85                  90                  95      


Cys Gly Lys Ala Gly Cys Gln Thr Tyr Lys Trp Glu Thr Leu Leu Thr 
            100                 105                 110         


Ser Glu Leu Pro Gln Trp Leu Ser Ala Asn Arg Ala Val Lys Pro Thr 
        115                 120                 125             


Gly Ser Ala Ala Ile Gly Leu Ser Met Ala Gly Ser Ser Ala Met Ile 
    130                 135                 140                 


Leu Ala Ala Tyr His Pro Gln Gln Phe Ile Tyr Ala Gly Ser Leu Ser 
145                 150                 155                 160 


Ala Leu Leu Asp Pro Ser Gln Gly Met Gly Pro Ser Leu Ile Gly Leu 
                165                 170                 175     


Ala Met Gly Asp Ala Gly Gly Tyr Lys Ala Ala Asp Met Trp Gly Pro 
            180                 185                 190         


Ser Ser Asp Pro Ala Trp Glu Arg Asn Asp Pro Thr Gln Gln Ile Pro 
        195                 200                 205             


Lys Leu Val Ala Asn Asn Thr Arg Leu Trp Val Tyr Cys Gly Asn Gly 
    210                 215                 220                 


Thr Pro Asn Glu Leu Gly Gly Ala Asn Ile Pro Ala Glu Phe Leu Glu 
225                 230                 235                 240 


Asn Phe Val Arg Ser Ser Asn Leu Lys Phe Gln Asp Ala Tyr Asn Ala 
                245                 250                 255     


Ala Gly Gly His Asn Ala Val Phe Asn Phe Pro Pro Asn Gly Thr His 
            260                 265                 270         


Ser Trp Glu Tyr Trp Gly Ala Gln Leu Asn Ala Met Lys Gly Asp Leu 
        275                 280                 285             


Gln Ser Ser Leu Gly Ala Gly Arg Ser 
    290                 295         


<210>  9
<211>  184
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Synthetic protein product 6X His-tag-Rv2660c::ESAT6

<400>  9

Met His His His His His His Gly Gly Ser Val Ile Ala Gly Val Asp 
1               5                   10                  15      


Gln Ala Leu Ala Ala Thr Gly Gln Ala Ser Gln Arg Ala Ala Gly Ala 
            20                  25                  30          


Ser Gly Gly Val Thr Val Gly Val Gly Val Gly Thr Glu Gln Arg Asn 
        35                  40                  45              


Leu Ser Val Val Ala Pro Ser Gln Phe Thr Phe Ser Ser Arg Ser Pro 
    50                  55                  60                  


Asp Phe Val Asp Glu Thr Ala Gly Gln Ser Trp Cys Ala Ile Leu Gly 
65                  70                  75                  80  


Leu Asn Gln Phe His Arg Ser Met Thr Glu Gln Gln Trp Asn Phe Ala 
                85                  90                  95      


Gly Ile Glu Ala Ala Ala Ser Ala Ile Gln Gly Asn Val Thr Ser Ile 
            100                 105                 110         


His Ser Leu Leu Asp Glu Gly Lys Gln Ser Leu Thr Lys Leu Ala Ala 
        115                 120                 125             


Ala Trp Gly Gly Ser Gly Ser Glu Ala Tyr Gln Gly Val Gln Gln Lys 
    130                 135                 140                 


Trp Asp Ala Thr Ala Thr Glu Leu Asn Asn Ala Leu Gln Asn Leu Ala 
145                 150                 155                 160 


Arg Thr Ile Ser Glu Ala Gly Gln Ala Met Ala Ser Thr Glu Gly Asn 
                165                 170                 175     


Val Thr Gly Met Phe Ala Arg Ser 
            180                 


<210>  10
<211>  42
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Synthetic primer

<400>  10
gggaagcttg ggtctagaag tttcgctcgc ggtgagtggc ga                          42


<210>  11
<211>  30
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Synthetic primer

<400>  11
gggactagtt caggccgaaa gtgccttgaa                                        30


