                         SEQUENCE LISTING

<110>  Lucite International UK Limited
 
<120>  Process for the Biological Production of Methacrylic Acid

<130>  P31059WO

<150>  GB1508582.2
<151>  2015-05-19

<150>  GB1517545.8
<151>  2015-10-05

<160>  24    

<170>  PatentIn version 3.5

<210>  1
<211>  471
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  optimised 4-hydroxybenzoyl-coA thioesterase (4HBT) from 
       Arthrobacter sp. strain SU

<400>  1
tatacatatg caccgtacct ctaacggttc tcacgcgacc ggtggtaacc tgccggatgt       60

tgcttctcat tacccggttg cgtacgaaca gaccctggac ggcaccgtgg gtttcgttat      120

cgacgaaatg accccggaac gtgcgaccgc gtctgttgaa gttaccgaca ccctgcgtca      180

gcgttggggt ctggttcacg gtggtgctta ctgcgcactg gcggagatgc tggcgaccga      240

agcgaccgtt gcggttgtgc acgaaaaggg tatgatggcg gttggccagt ctaaccacac      300

ctctttcttc cgtccggtta aagaaggtca cgttcgcgcg gaagctgttc gcatccacgc      360

gggttctacc acctggttct gggacgtttc cctgcgcgat gatgcgggtc gcctgtgtgc      420

ggtgtcttct atgtctatcg cggtgcgtcc gcgtcgtgac taagcggccg c               471


<210>  2
<211>  36
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  HHT.F forward primer for cloning of 4-HBT

<400>  2
tatacatatg caccgtacct ctaacggttc tcacgc                                 36


<210>  3
<211>  24
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  HHT.R reverse primer for cloning of 4HBT

<400>  3
ctcgagtccg tcacgacgcg gacg                                              24


<210>  4
<211>  469
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Product of the PCR performed to modify the 4HBT gene for sub 
       cloning into the pET20b(+) plasmid to form the pET20b(+)::CtHis 
       4HBT plasmid.

<400>  4
tatacatatg caccgtacct ctaacggttc tcacgcgacc ggtggtaacc tgccggatgt       60

tgcttctcat tacccggttg cgtacgaaca gaccctggac ggcaccgtgg gtttcgttat      120

cgacgaaatg accccggaac gtgcgaccgc gtctgttgaa gttaccgaca ccctgcgtca      180

gcgttggggt ctggttcacg gtggtgctta ctgcgcactg gcggagatgc tggcgaccga      240

agcgaccgtt gcggttgtgc acgaaaaggg tatgatggcg gttggccagt ctaaccacac      300

ctctttcttc cgtccggtta aagaaggtca cgttcgcgcg gaagctgttc gcatccacgc      360

gggttctacc acctggttct gggacgtttc cctgcgcgat gatgcgggtc gcctgtgtgc      420

ggtgtcttct atgtctatcg cggtgcgtcc gcgtcgtgac ggactcgag                  469


<210>  5
<211>  483
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  carboxy-terminal hexahistidine tagged 4HBT in the 
       pET20b(+)::CtHis-4HBT plasmid

<400>  5
atgcaccgta cctctaacgg ttctcacgcg accggtggta acctgccgga tgttgcttct       60

cattacccgg ttgcgtacga acagaccctg gacggcaccg tgggtttcgt tatcgacgaa      120

atgaccccgg aacgtgcgac cgcgtctgtt gaagttaccg acaccctgcg tcagcgttgg      180

ggtctggttc acggtggtgc ttactgcgca ctggcggaga tgctggcgac cgaagcgacc      240

gttgcggttg tgcacgaaaa gggtatgatg gcggttggcc agtctaacca cacctctttc      300

ttccgtccgg ttaaagaagg tcacgttcgc gcggaagctg ttcgcatcca cgcgggttct      360

accacctggt tctgggacgt ttccctgcgc gatgatgcgg gtcgcctgtg tgcggtgtct      420

tctatgtcta tcgcggtgcg tccgcgtcgt gacggactcg agcaccacca ccaccaccac      480

tga                                                                    483


<210>  6
<211>  1320
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  optimised acyl-coA oxidase 4 (ACX4) from Arabidopsis thaliana

<400>  6
catatggcag ttctgagcag cgcagatcgt gcaagcaatg aaaaaaaagt gaaaagcagc       60

tatttcgacc tgcctccgat ggaaatgagc gttgcatttc cgcaggcaac accggcaagc      120

acctttccgc cttgtaccag cgattattat cattttaatg atctgctgac ccctgaagaa      180

caggccattc gtaaaaaagt tcgtgagtgc atggaaaaag aagtggcacc gattatgacc      240

gaatattggg aaaaagcaga attcccgttt catattaccc cgaaactggg tgcaatgggt      300

gttgccggtg gtagcattaa aggttatggt tgtccgggtc tgagcattac cgcaaatgca      360

attgcaaccg cagaaattgc acgtgttgat gcaagctgta gcacctttat tctggttcat      420

agcagcctgg gtatgctgac cattgcactg tgtggtagcg aagcacagaa agaaaaatat      480

ctgccgagcc tggcacagct gaataccgtt gcatgttggg cactgaccga accggataat      540

ggtagtgatg caagcggtct gggcaccacc gcaaccaaag ttgaaggtgg ttggaaaatt      600

aacggtcaga aacgttggat tggcaatagc acctttgcag atctgctgat tatctttgca      660

cgtaatacca ccaccaatca gattaacggc ttcatcgtta aaaaagatgc accgggtctg      720

aaagcaacca aaattccgaa caaaattggt ctgcgtatgg tgcagaatgg tgatattctg      780

ctgcagaatg tttttgtgcc ggatgaagat cgtctgcctg gtgttaatag ctttcaggat      840

accagcaaag ttctggcagt tagccgtgtt atggttgcat ggcagccgat tggtattagc      900

atgggtattt atgatatgtg ccaccgctat ctgaaagaac gtaaacagtt tggtgcaccg      960

ctggcagcat ttcagctgaa tcagcagaaa ctggttcaga tgctgggtaa tgttcaggca     1020

atgtttctga tgggttggcg tctgtgtaaa ctgtatgaaa ccggtcagat gacaccgggt     1080

caggcaagcc tgggtaaagc atggattagc agcaaagcac gtgaaaccgc cagcctgggt     1140

cgtgaactgc tgggtggtaa tggtattctg gcagattttc tggttgcaaa agcattttgt     1200

gatctggaac cgatctatac ctatgaaggc acctatgata tcaataccct ggttaccggt     1260

cgtgaagtta ccggtattgc aagctttaaa ccggcaaccc gtagccgtct gtaactcgag     1320


<210>  7
<211>  27
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  OE.A.F forward primer for overlap extension polymerase chain 
       reaction A

<400>  7
acatatgcac cgtacctcta acggttc                                           27


<210>  8
<211>  39
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  OE.A.R reverse primer for overlap extension polymerisation chain 
       reaction B

<400>  8
ctgccatatc tatatctcct gttagtcacg acgcggacg                              39


<210>  9
<211>  40
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  OE.B.F forward primer for overlap extension polymerase chain 
       reaction B

<400>  9
gtgactaaca ggagatatag atatggcagt tctgagcagc                             40


<210>  10
<211>  40
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  OE.B.R reverse primer for overlap polymerase chain reaction B

<400>  10
ctcgagatat tatagctagc ttacagacgg ctacgggttg                             40


<210>  11
<211>  1805
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  product of the overlap polymerase chain reaction to concatenate 
       4HBT and ACX4 into one polynucleotide

<400>  11
acatatgcac cgtacctcta acggttctca cgcgaccggt ggtaacctgc cggatgttgc       60

ttctcattac ccggttgcgt acgaacagac cctggacggc accgtgggtt tcgttatcga      120

cgaaatgacc ccggaacgtg cgaccgcgtc tgttgaagtt accgacaccc tgcgtcagcg      180

ttggggtctg gttcacggtg gtgcttactg cgcactggcg gagatgctgg cgaccgaagc      240

gaccgttgcg gttgtgcacg aaaagggtat gatggcggtt ggccagtcta accacacctc      300

tttcttccgt ccggttaaag aaggtcacgt tcgcgcggaa gctgttcgca tccacgcggg      360

ttctaccacc tggttctggg acgtttccct gcgcgatgat gcgggtcgcc tgtgtgcggt      420

gtcttctatg tctatcgcgg tgcgtccgcg tcgtgactaa caggagatat agatatggca      480

gttctgagca gcgcagatcg tgcaagcaat gaaaaaaaag tgaaaagcag ctatttcgac      540

ctgcctccga tggaaatgag cgttgcattt ccgcaggcaa caccggcaag cacctttccg      600

ccttgtacca gcgattatta tcattttaat gatctgctga cccctgaaga acaggccatt      660

cgtaaaaaag ttcgtgagtg catggaaaaa gaagtggcac cgattatgac cgaatattgg      720

gaaaaagcag aattcccgtt tcatattacc ccgaaactgg gtgcaatggg tgttgccggt      780

ggtagcatta aaggttatgg ttgtccgggt ctgagcatta ccgcaaatgc aattgcaacc      840

gcagaaattg cacgtgttga tgcaagctgt agcaccttta ttctggttca tagcagcctg      900

ggtatgctga ccattgcact gtgtggtagc gaagcacaga aagaaaaata tctgccgagc      960

ctggcacagc tgaataccgt tgcatgttgg gcactgaccg aaccggataa tggtagtgat     1020

gcaagcggtc tgggcaccac cgcaaccaaa gttgaaggtg gttggaaaat taacggtcag     1080

aaacgttgga ttggcaatag cacctttgca gatctgctga ttatctttgc acgtaatacc     1140

accaccaatc agattaacgg cttcatcgtt aaaaaagatg caccgggtct gaaagcaacc     1200

aaaattccga acaaaattgg tctgcgtatg gtgcagaatg gtgatattct gctgcagaat     1260

gtttttgtgc cggatgaaga tcgtctgcct ggtgttaata gctttcagga taccagcaaa     1320

gttctggcag ttagccgtgt tatggttgca tggcagccga ttggtattag catgggtatt     1380

tatgatatgt gccaccgcta tctgaaagaa cgtaaacagt ttggtgcacc gctggcagca     1440

tttcagctga atcagcagaa actggttcag atgctgggta atgttcaggc aatgtttctg     1500

atgggttggc gtctgtgtaa actgtatgaa accggtcaga tgacaccggg tcaggcaagc     1560

ctgggtaaag catggattag cagcaaagca cgtgaaaccg ccagcctggg tcgtgaactg     1620

ctgggtggta atggtattct ggcagatttt ctggttgcaa aagcattttg tgatctggaa     1680

ccgatctata cctatgaagg cacctatgat atcaataccc tggttaccgg tcgtgaagtt     1740

accggtattg caagctttaa accggcaacc cgtagccgtc tgtaagctag ctataatatc     1800

tcgag                                                                 1805


<210>  12
<211>  1663
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  optimised acyl-CoA synthetase (acsA) from Pseudomonas 
       chlororaphis B23

<400>  12
gctagcagga gatatacata tgcgcgacta cgaacacgtt gtggaatctt tcgactacct       60

gcagagcgca acccaggacc tgcatggcga actgactgct ctgaacgcgt gtgtggagtg      120

ctgtgatcgt cacgcgcacg gcgaggctgt tgctctgtat tgcgaagcgc aggatggcca      180

cgcggaacgc taccgtttcc gcgatctgca gcgccaggct gcacgttttg gcaacttcct      240

gcgtgagcag ggcgtgaaac cgggcgaccg tgttgcgggc ctgatgccgc gtaccgtgga      300

actgctgatc gcaatcctgg gcacttggcg catcggtgcg gtttatcagc cgctgtttac      360

tgcgttcggt ccgaaagcaa ttgagcagcg tctgaactgt tctaacgcac gctggattgt      420

gaccgatccg cacaaccgcc cgaaactgga cgacgttacc gactgcccgt ctattgtggt      480

taccggtggc gcaccgcaga acccggcaga ccatcacttt tggtccgcac tgaaccgtca      540

ggcggatgac tgtgcgccgg tgctgctgga cgcatctgca ccgttcctgc tgatgtgcac      600

ctccggcacc actggtccgg caaagccgct ggaagttccg ctgtctgcga ttctggcatt      660

taaaggttac atgcgtgatg caatcgacct gcgcgctgat gatcgcttct ggaacctggc      720

agatccgggt tgggcatacg gtctgtacta cgcggtgact ggcccgctgg cgtgtggcta      780

cgcgaccctg ttttacgacg gtccgttcac tgttgaatcc acccgtcaca tcatcgcgaa      840

atacgcgatt aacaacctgg cgggtagccc gaccgcttac cgctttctga ttgctgcagg      900

cgcggaattt gcggacgctg ttcgtggtcg tctgcgtgca gttagcagcg cgggtgaacc      960

gctgaacccg caggtggttc gttggtttgc tgaacagctg ggtgttgtta tccatgacca     1020

ctatggtcag accgaaattg gtatggttct gtgcaaccac cacggcctgc gtcacccggt     1080

tcgcgagggt agcgctggct atgcagtgcc gggctatcgt atcgtggtgc tggataaagc     1140

acaccgtgag ctgccggctg gtcagccggg tgttctggcg gtggaccgtg aacgctcccc     1200

gctgtgctgg tttgacggct acctgggtat gccgactcag gcgttcgcgg gtcgttatta     1260

cctgtctggt gatatcgttg aactgaacga cgatggcagc atctctttcg tgggtcgcaa     1320

cgatgatctg atcaccacct ctggttatcg tgtgggcccg ttcgacgttg agtctgcact     1380

gattgaacat ccggcagttg ttgaagctgc tgttatcggt aaaccggacc cgcagcgtac     1440

tgagctgatt aaagcgtttg ttgttctgaa cactccgtac ctgccgagcc cggaactggc     1500

ggaagaactg cgtctgcatg ttcgtcagcg cctggctgcg cacgcatatc cgcgcgagat     1560

ggagtttgtg gaccatctgc cgaaaacccc gtctggcaaa ctgcagcgtt tcattctgcg     1620

taaccaggaa atcgctaagc agcaggctct gggttaactc gag                       1663


<210>  13
<211>  3447
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  The sequence between, and inclusive of, the NdeI and XhoI 
       restriction sites in pET20b(+)::4HBT ACX4 AcsA.

<400>  13
catatgcacc gtacctctaa cggttctcac gcgaccggtg gtaacctgcc ggatgttgct       60

tctcattacc cggttgcgta cgaacagacc ctggacggca ccgtgggttt cgttatcgac      120

gaaatgaccc cggaacgtgc gaccgcgtct gttgaagtta ccgacaccct gcgtcagcgt      180

tggggtctgg ttcacggtgg tgcttactgc gcactggcgg agatgctggc gaccgaagcg      240

accgttgcgg ttgtgcacga aaagggtatg atggcggttg gccagtctaa ccacacctct      300

ttcttccgtc cggttaaaga aggtcacgtt cgcgcggaag ctgttcgcat ccacgcgggt      360

tctaccacct ggttctggga cgtttccctg cgcgatgatg cgggtcgcct gtgtgcggtg      420

tcttctatgt ctatcgcggt gcgtccgcgt cgtgactaac aggagatata gatatggcag      480

ttctgagcag cgcagatcgt gcaagcaatg aaaaaaaagt gaaaagcagc tatttcgacc      540

tgcctccgat ggaaatgagc gttgcatttc cgcaggcaac accggcaagc acctttccgc      600

cttgtaccag cgattattat cattttaatg atctgctgac ccctgaagaa caggccattc      660

gtaaaaaagt tcgtgagtgc atggaaaaag aagtggcacc gattatgacc gaatattggg      720

aaaaagcaga attcccgttt catattaccc cgaaactggg tgcaatgggt gttgccggtg      780

gtagcattaa aggttatggt tgtccgggtc tgagcattac cgcaaatgca attgcaaccg      840

cagaaattgc acgtgttgat gcaagctgta gcacctttat tctggttcat agcagcctgg      900

gtatgctgac cattgcactg tgtggtagcg aagcacagaa agaaaaatat ctgccgagcc      960

tggcacagct gaataccgtt gcatgttggg cactgaccga accggataat ggtagtgatg     1020

caagcggtct gggcaccacc gcaaccaaag ttgaaggtgg ttggaaaatt aacggtcaga     1080

aacgttggat tggcaatagc acctttgcag atctgctgat tatctttgca cgtaatacca     1140

ccaccaatca gattaacggc ttcatcgtta aaaaagatgc accgggtctg aaagcaacca     1200

aaattccgaa caaaattggt ctgcgtatgg tgcagaatgg tgatattctg ctgcagaatg     1260

tttttgtgcc ggatgaagat cgtctgcctg gtgttaatag ctttcaggat accagcaaag     1320

ttctggcagt tagccgtgtt atggttgcat ggcagccgat tggtattagc atgggtattt     1380

atgatatgtg ccaccgctat ctgaaagaac gtaaacagtt tggtgcaccg ctggcagcat     1440

ttcagctgaa tcagcagaaa ctggttcaga tgctgggtaa tgttcaggca atgtttctga     1500

tgggttggcg tctgtgtaaa ctgtatgaaa ccggtcagat gacaccgggt caggcaagcc     1560

tgggtaaagc atggattagc agcaaagcac gtgaaaccgc cagcctgggt cgtgaactgc     1620

tgggtggtaa tggtattctg gcagattttc tggttgcaaa agcattttgt gatctggaac     1680

cgatctatac ctatgaaggc acctatgata tcaataccct ggttaccggt cgtgaagtta     1740

ccggtattgc aagctttaaa ccggcaaccc gtagccgtct gtaagctagc aggagatata     1800

catatgcgcg actacgaaca cgttgtggaa tctttcgact acctgcagag cgcaacccag     1860

gacctgcatg gcgaactgac tgctctgaac gcgtgtgtgg agtgctgtga tcgtcacgcg     1920

cacggcgagg ctgttgctct gtattgcgaa gcgcaggatg gccacgcgga acgctaccgt     1980

ttccgcgatc tgcagcgcca ggctgcacgt tttggcaact tcctgcgtga gcagggcgtg     2040

aaaccgggcg accgtgttgc gggcctgatg ccgcgtaccg tggaactgct gatcgcaatc     2100

ctgggcactt ggcgcatcgg tgcggtttat cagccgctgt ttactgcgtt cggtccgaaa     2160

gcaattgagc agcgtctgaa ctgttctaac gcacgctgga ttgtgaccga tccgcacaac     2220

cgcccgaaac tggacgacgt taccgactgc ccgtctattg tggttaccgg tggcgcaccg     2280

cagaacccgg cagaccatca cttttggtcc gcactgaacc gtcaggcgga tgactgtgcg     2340

ccggtgctgc tggacgcatc tgcaccgttc ctgctgatgt gcacctccgg caccactggt     2400

ccggcaaagc cgctggaagt tccgctgtct gcgattctgg catttaaagg ttacatgcgt     2460

gatgcaatcg acctgcgcgc tgatgatcgc ttctggaacc tggcagatcc gggttgggca     2520

tacggtctgt actacgcggt gactggcccg ctggcgtgtg gctacgcgac cctgttttac     2580

gacggtccgt tcactgttga atccacccgt cacatcatcg cgaaatacgc gattaacaac     2640

ctggcgggta gcccgaccgc ttaccgcttt ctgattgctg caggcgcgga atttgcggac     2700

gctgttcgtg gtcgtctgcg tgcagttagc agcgcgggtg aaccgctgaa cccgcaggtg     2760

gttcgttggt ttgctgaaca gctgggtgtt gttatccatg accactatgg tcagaccgaa     2820

attggtatgg ttctgtgcaa ccaccacggc ctgcgtcacc cggttcgcga gggtagcgct     2880

ggctatgcag tgccgggcta tcgtatcgtg gtgctggata aagcacaccg tgagctgccg     2940

gctggtcagc cgggtgttct ggcggtggac cgtgaacgct ccccgctgtg ctggtttgac     3000

ggctacctgg gtatgccgac tcaggcgttc gcgggtcgtt attacctgtc tggtgatatc     3060

gttgaactga acgacgatgg cagcatctct ttcgtgggtc gcaacgatga tctgatcacc     3120

acctctggtt atcgtgtggg cccgttcgac gttgagtctg cactgattga acatccggca     3180

gttgttgaag ctgctgttat cggtaaaccg gacccgcagc gtactgagct gattaaagcg     3240

tttgttgttc tgaacactcc gtacctgccg agcccggaac tggcggaaga actgcgtctg     3300

catgttcgtc agcgcctggc tgcgcacgca tatccgcgcg agatggagtt tgtggaccat     3360

ctgccgaaaa ccccgtctgg caaactgcag cgtttcattc tgcgtaacca ggaaatcgct     3420

aagcagcagg ctctgggtta actcgag                                         3447


<210>  14
<211>  4996
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  ppBCKD polynucleotide containing the four genes encoding the 
       subunits of the Pseudomonas putida KT2440 branched chain keto 
       acid dehydrogenase delivered in the pBSK::ppBCKD plasmid

<400>  14
tctagaaata attttgttta actgctagca ggagaaatta actatgaacg agtacgcccc       60

cctgcgtttg catgtgcccg agcccaccgg ccggccaggc tgccagaccg atttttccta      120

cctgcgcctc aacgatgcag gtcaagcccg taaacccgcg atcgatgtcg atgctgccga      180

cactgccgac ctgtcctaca gcctggtccg cgtgctcgac gagcaaggcg atgcgcaagg      240

cccctgggcc gaagacatcg acccgcagat cctccgtcaa ggcatgcgcg ccatgctcaa      300

gacgcggatc ttcgacagcc gcatggtggt tgcccagcgc cagaagaaga tgtccttcta      360

catgcaaagc ctgggcgaag aagccatcgg cagcggccag gcgctggcgc tgaaccgcac      420

cgacatgtgc ttcccgacct accgccagca aagcatcctg atggcccgcg acgtgtcgct      480

ggtcgagatg atctgccaac tgctgtccaa cgagcgcgac cccctcaagg gccgccagtt      540

gccgatcatg tattcggtgc gcgaagccgg cttcttcacc atcagcggca acctggcgac      600

ccagttcgtg caggcggtcg gctgggccat ggcatcggcg atcaagggcg ataccaagat      660

cgcctcggca tggatcggtg acggcgctac tgccgagtcg gacttccaca ccgcccttac      720

ctttgcccac gtataccgcg ccccggtcat cctcaacgtg gtcaacaacc aatgggccat      780

ttccaccttc caggccatcg ccggtggcga gtcgaccacc tttgccggcc gtggcgtggg      840

ttgcggtatt gcttcgctgc gggttgacgg caacgacttc gtcgccgtgt acgctgcctc      900

gcgctgggcg gccgagcgcg cccgccgcgg cctgggccca agcctgatcg agtgggtcac      960

ctaccgtgcc ggcccgcact cgacgtcgga cgacccctcc aagtaccgcc ctgccgacga     1020

ctggagccac ttcccgctgg gtgacccgat cgcccgcctg aagcagcacc tgatcaagat     1080

cggccactgg tccgaggaag aacaccaggc cgtcacggcc gagctcgaag ctgcggtgat     1140

tgccgcacag aaagaagccg agcagtacgg caccctggcc aacgggcaca tcccgagcgc     1200

cgcctcgatg ttcgaggatg tgtacaagga aatgcccgac cacctgcgcc gtcaacgcca     1260

ggaactgggg gtttgagatg aacgaccaca acaacagcat caacccggaa accgccatgg     1320

ccaccactac catgaccatg atccaggccc tgcgctcggc catggatgtc atgcttgagc     1380

gcgacgacaa tgtggtggtg tacggccagg acgtcggtta cttcggcggc gtgttccgct     1440

gcaccgaagg cctgcagaac aagtacggca aatcgcgcgt gttcgacgcg cccatctccg     1500

agagcggcat cgtcggtacc gccgtgggca tgggtgccta tggcctgcgc ccggtggtgg     1560

agatccagtt cgccgactac ttctacccgg cctccgacca gatcgtctcc gagctggccc     1620

gcctgcgtta ccgttcggcc ggcgagttca ttgccccgct gaccctgcgc atgccttgcg     1680

gcggcggcat ctatggcggc cagactcaca gccagagccc ggaagcgatg ttcacccagg     1740

tgtgcggcct gcgcaccgtg atgccgtcca acccttatga cgccaaaggc ctgttgattg     1800

cctcgatcga atgcgacgac ccggtaatct tcctggagcc caaacgcctg tacaacggcc     1860

cgttcgatgg ccaccacgac cgccctgtaa ccccgtggtc gaagcacccg cacagcgccg     1920

tgcccgacgg ttattacacc gtaccgctgg acaaggccgc cattacccgc cctggcaatg     1980

acgtgaccgt gctgacctac ggcaccacgg tgtacgtggc ccaggtggcc gccgaagaaa     2040

gcggcgtcga tgccgaagtg atcgacctgc gcagcctgtg gccgctggac ctggacacta     2100

tcgtcgagtc ggtgaaaaag accggccgtt gcgtggtggt gcacgaggcc acccgcacct     2160

gcggcttcgg tgccgagctg gtgtcgctgg tgcaggagca ctgcttccac cacctggagg     2220

cgccgatcga acgcgtcacc ggctgggaca ccccctaccc tcacgcacag gaatgggctt     2280

acttcccagg cccttcgcgg gtaggtgcgg cactgaaaaa ggtcatggag gtctgaatgg     2340

gcacgcacgt catcaagatg ccggacattg gcgaaggcat cgcgcaggtc gagttggtgg     2400

aatggttcgt caaggtcggc gacatcatcg ccgaggacca ggtggtggcc gacgtcatga     2460

ccgacaaggc caccgtggaa atcccctcgc cggtcagcgg caaggtgttg gccctgggtg     2520

gccagcccgg ggaagtgatg gcggtcggta gcgaactgat ccgcatcgaa gtggaaggca     2580

gcggcaacca tgtggatgtg cctcagccaa aaccggtaga ggccccggct gcccccattg     2640

cagccaagcc ggaaccgcag aaagacgtaa aacccgccgt gtaccaggcg cccgccaacc     2700

acgaagctgc gcccatcgtg ccgcgccagc cgggcgacaa gccgctggcc tcgcctgccg     2760

tgcgcaaacg cgccctggac gccggtatcg aactgcgtta tgtgcatggt agcggcccgg     2820

ccgggcgtat tctgcacgaa gacctcgatg ccttcatgag caagccgcaa agcaatgccg     2880

ggcaagcacc tgatggttat gccaagcgca ccgacagcga gcaggtgcca gtgatcggcc     2940

tgcgccgcaa gattgcccag cgcatgcagg acgccaaacg ccgggtcgcg cacttcagtt     3000

atgtcgagga aatcgacgtc accgccctgg aggccctgcg ccagcaactc aacagcaagc     3060

acggcgacag ccggggcaag ctgaccttgc tgccattcct ggtacgcgcc ctggtcgtgg     3120

cgctgcgtga cttcccgcag atcaacgcga cctacgatga cgaagcgcag atcatcaccc     3180

gccatggcgc ggtgcatgtg ggcattgcca cccaaggtga caacggcctg atggtgcccg     3240

tgctgcgcca cgccgaagcg ggcagcctgt gggccaatgc cggcgagatt tcgcgcctgg     3300

ccaacgctgc acgtaacaac aaggccagcc gtgaagagct gtccggctcg accatcaccc     3360

tgaccagcct tggcgccctg ggtggcattg tcagcacgcc ggtggtcaac accccggaag     3420

tggcaatcgt cggggtcaac cgcatggtcg aacggccagt ggtgatcgac ggccagatcg     3480

tcgtgcgcaa gatgatgaac ctgtccagct cgttcgacca ccgcgtggtc gatggcatgg     3540

atgccgccct gttcatccag gccgtacgtg gcctgctcga acaacccgcc tgcctgttcg     3600

tggagtgagc atgcaacaga ttatccagac taccctgttg atcatcggcg gcggccctgg     3660

cggctatgta gcagccatcc gcgccgggca actgggcatt cctaccgtac tggtggaagg     3720

ccaggcactg ggcggcacct gcctgaacat cggctgcatc ccgtccaagg cgctgatcca     3780

cgtggccgag cagtttcacc aggcctcgcg ctttaccgaa ccctcgccgc tgggcatcag     3840

cgtggcttcg ccgcgcctgg acatcggcca gagcgtcacc tggaaggacg gcattgtcga     3900

ccgcctgacc acaggtgttg ccgccctgct gaaaaagcac ggggtgaaag tggtgcatgg     3960

ttgggccaag gtactggacg gcaagcaggt cgaggtcgat ggccagcgta tccagtgcga     4020

gcatctgttg ctggcgaccg gttccagcag tgtcgaactg cctatgctgc cgctgggtgg     4080

cccggtgatt tcctcgaccg aagccctggc gccgaaaacc ctgccgcaac acctggtggt     4140

ggtcggcggt ggctatatcg gcctggagct gggcattgcc tatcgcaagc tgggtgcaca     4200

ggtgagtgtg gtggaagcgc gggagcgcat cctgccgacc tacgacagcg aattgaccgc     4260

cccggtggcc gagtcgctga agaaactggg catagcgttg cacctgggcc acagcgtcga     4320

gggctacgaa aatggctgcc tgctggccag cgacggcaag ggtgggcaac tgcgccttga     4380

ggccgaccag gtactggtgg ccgtgggacg ccggccacgc accaagggct tcaacctgga     4440

atgcctggac ctgaagatga acggcgccgc cattgccatc gacgagcgct gtcacaccag     4500

catgcacaac gtctgggcca tcggcgacgt cgctggcgaa ccgatgctgg cgcaccgggc     4560

catggcccag ggcgagatgg tcgccgaaat catcgccggc aaggcccgac gctttgaacc     4620

gacagcgatt gccgccgtgt gctttaccga cccggaagtg gtggtggtcg gcaagacccc     4680

ggaacaagcc agccagcagg gcctggactg catcgtcgcg cagttcccgt ttgccgccaa     4740

tggccgggcc atgagcctgg aatcgaaaag cggtttcgtg cgggtggtgg cgcgccgtga     4800

caaccacctg atcgtgggtt ggcaggcggt tggcgtggcg gtctccgagc tatccaccgc     4860

gtttgcccaa tcgctggaga tgggcgcgtg cctggaagat gtggccggta ccattcatgc     4920

ccacccaacg ctcggtgaag cggtacagga agccgcactg cgcgccctgg gccacgcctt     4980

gcatatctga ctcgag                                                     4996


<210>  15
<211>  6758
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  The sequence between, and inclusive of, the NdeI and XhoI 
       restriction sites in the pET20b(+)::4HBT ACX4 ppBCKD plasmid

<400>  15
acatatgcac cgtacctcta acggttctca cgcgaccggt ggtaacctgc cggatgttgc       60

ttctcattac ccggttgcgt acgaacagac cctggacggc accgtgggtt tcgttatcga      120

cgaaatgacc ccggaacgtg cgaccgcgtc tgttgaagtt accgacaccc tgcgtcagcg      180

ttggggtctg gttcacggtg gtgcttactg cgcactggcg gagatgctgg cgaccgaagc      240

gaccgttgcg gttgtgcacg aaaagggtat gatggcggtt ggccagtcta accacacctc      300

tttcttccgt ccggttaaag aaggtcacgt tcgcgcggaa gctgttcgca tccacgcggg      360

ttctaccacc tggttctggg acgtttccct gcgcgatgat gcgggtcgcc tgtgtgcggt      420

gtcttctatg tctatcgcgg tgcgtccgcg tcgtgactaa caggagatat agatatggca      480

gttctgagca gcgcagatcg tgcaagcaat gaaaaaaaag tgaaaagcag ctatttcgac      540

ctgcctccga tggaaatgag cgttgcattt ccgcaggcaa caccggcaag cacctttccg      600

ccttgtacca gcgattatta tcattttaat gatctgctga cccctgaaga acaggccatt      660

cgtaaaaaag ttcgtgagtg catggaaaaa gaagtggcac cgattatgac cgaatattgg      720

gaaaaagcag aattcccgtt tcatattacc ccgaaactgg gtgcaatggg tgttgccggt      780

ggtagcatta aaggttatgg ttgtccgggt ctgagcatta ccgcaaatgc aattgcaacc      840

gcagaaattg cacgtgttga tgcaagctgt agcaccttta ttctggttca tagcagcctg      900

ggtatgctga ccattgcact gtgtggtagc gaagcacaga aagaaaaata tctgccgagc      960

ctggcacagc tgaataccgt tgcatgttgg gcactgaccg aaccggataa tggtagtgat     1020

gcaagcggtc tgggcaccac cgcaaccaaa gttgaaggtg gttggaaaat taacggtcag     1080

aaacgttgga ttggcaatag cacctttgca gatctgctga ttatctttgc acgtaatacc     1140

accaccaatc agattaacgg cttcatcgtt aaaaaagatg caccgggtct gaaagcaacc     1200

aaaattccga acaaaattgg tctgcgtatg gtgcagaatg gtgatattct gctgcagaat     1260

gtttttgtgc cggatgaaga tcgtctgcct ggtgttaata gctttcagga taccagcaaa     1320

gttctggcag ttagccgtgt tatggttgca tggcagccga ttggtattag catgggtatt     1380

tatgatatgt gccaccgcta tctgaaagaa cgtaaacagt ttggtgcacc gctggcagca     1440

tttcagctga atcagcagaa actggttcag atgctgggta atgttcaggc aatgtttctg     1500

atgggttggc gtctgtgtaa actgtatgaa accggtcaga tgacaccggg tcaggcaagc     1560

ctgggtaaag catggattag cagcaaagca cgtgaaaccg ccagcctggg tcgtgaactg     1620

ctgggtggta atggtattct ggcagatttt ctggttgcaa aagcattttg tgatctggaa     1680

ccgatctata cctatgaagg cacctatgat atcaataccc tggttaccgg tcgtgaagtt     1740

accggtattg caagctttaa accggcaacc cgtagccgtc tgtaagctag caggagaaat     1800

taactatgaa cgagtacgcc cccctgcgtt tgcatgtgcc cgagcccacc ggccggccag     1860

gctgccagac cgatttttcc tacctgcgcc tcaacgatgc aggtcaagcc cgtaaacccg     1920

cgatcgatgt cgatgctgcc gacactgccg acctgtccta cagcctggtc cgcgtgctcg     1980

acgagcaagg cgatgcgcaa ggcccctggg ccgaagacat cgacccgcag atcctccgtc     2040

aaggcatgcg cgccatgctc aagacgcgga tcttcgacag ccgcatggtg gttgcccagc     2100

gccagaagaa gatgtccttc tacatgcaaa gcctgggcga agaagccatc ggcagcggcc     2160

aggcgctggc gctgaaccgc accgacatgt gcttcccgac ctaccgccag caaagcatcc     2220

tgatggcccg cgacgtgtcg ctggtcgaga tgatctgcca actgctgtcc aacgagcgcg     2280

accccctcaa gggccgccag ttgccgatca tgtattcggt gcgcgaagcc ggcttcttca     2340

ccatcagcgg caacctggcg acccagttcg tgcaggcggt cggctgggcc atggcatcgg     2400

cgatcaaggg cgataccaag atcgcctcgg catggatcgg tgacggcgct actgccgagt     2460

cggacttcca caccgccctt acctttgccc acgtataccg cgccccggtc atcctcaacg     2520

tggtcaacaa ccaatgggcc atttccacct tccaggccat cgccggtggc gagtcgacca     2580

cctttgccgg ccgtggcgtg ggttgcggta ttgcttcgct gcgggttgac ggcaacgact     2640

tcgtcgccgt gtacgctgcc tcgcgctggg cggccgagcg cgcccgccgc ggcctgggcc     2700

caagcctgat cgagtgggtc acctaccgtg ccggcccgca ctcgacgtcg gacgacccct     2760

ccaagtaccg ccctgccgac gactggagcc acttcccgct gggtgacccg atcgcccgcc     2820

tgaagcagca cctgatcaag atcggccact ggtccgagga agaacaccag gccgtcacgg     2880

ccgagctcga agctgcggtg attgccgcac agaaagaagc cgagcagtac ggcaccctgg     2940

ccaacgggca catcccgagc gccgcctcga tgttcgagga tgtgtacaag gaaatgcccg     3000

accacctgcg ccgtcaacgc caggaactgg gggtttgaga tgaacgacca caacaacagc     3060

atcaacccgg aaaccgccat ggccaccact accatgacca tgatccaggc cctgcgctcg     3120

gccatggatg tcatgcttga gcgcgacgac aatgtggtgg tgtacggcca ggacgtcggt     3180

tacttcggcg gcgtgttccg ctgcaccgaa ggcctgcaga acaagtacgg caaatcgcgc     3240

gtgttcgacg cgcccatctc cgagagcggc atcgtcggta ccgccgtggg catgggtgcc     3300

tatggcctgc gcccggtggt ggagatccag ttcgccgact acttctaccc ggcctccgac     3360

cagatcgtct ccgagctggc ccgcctgcgt taccgttcgg ccggcgagtt cattgccccg     3420

ctgaccctgc gcatgccttg cggcggcggc atctatggcg gccagactca cagccagagc     3480

ccggaagcga tgttcaccca ggtgtgcggc ctgcgcaccg tgatgccgtc caacccttat     3540

gacgccaaag gcctgttgat tgcctcgatc gaatgcgacg acccggtaat cttcctggag     3600

cccaaacgcc tgtacaacgg cccgttcgat ggccaccacg accgccctgt aaccccgtgg     3660

tcgaagcacc cgcacagcgc cgtgcccgac ggttattaca ccgtaccgct ggacaaggcc     3720

gccattaccc gccctggcaa tgacgtgacc gtgctgacct acggcaccac ggtgtacgtg     3780

gcccaggtgg ccgccgaaga aagcggcgtc gatgccgaag tgatcgacct gcgcagcctg     3840

tggccgctgg acctggacac tatcgtcgag tcggtgaaaa agaccggccg ttgcgtggtg     3900

gtgcacgagg ccacccgcac ctgcggcttc ggtgccgagc tggtgtcgct ggtgcaggag     3960

cactgcttcc accacctgga ggcgccgatc gaacgcgtca ccggctggga caccccctac     4020

cctcacgcac aggaatgggc ttacttccca ggcccttcgc gggtaggtgc ggcactgaaa     4080

aaggtcatgg aggtctgaat gggcacgcac gtcatcaaga tgccggacat tggcgaaggc     4140

atcgcgcagg tcgagttggt ggaatggttc gtcaaggtcg gcgacatcat cgccgaggac     4200

caggtggtgg ccgacgtcat gaccgacaag gccaccgtgg aaatcccctc gccggtcagc     4260

ggcaaggtgt tggccctggg tggccagccc ggggaagtga tggcggtcgg tagcgaactg     4320

atccgcatcg aagtggaagg cagcggcaac catgtggatg tgcctcagcc aaaaccggta     4380

gaggccccgg ctgcccccat tgcagccaag ccggaaccgc agaaagacgt aaaacccgcc     4440

gtgtaccagg cgcccgccaa ccacgaagct gcgcccatcg tgccgcgcca gccgggcgac     4500

aagccgctgg cctcgcctgc cgtgcgcaaa cgcgccctgg acgccggtat cgaactgcgt     4560

tatgtgcatg gtagcggccc ggccgggcgt attctgcacg aagacctcga tgccttcatg     4620

agcaagccgc aaagcaatgc cgggcaagca cctgatggtt atgccaagcg caccgacagc     4680

gagcaggtgc cagtgatcgg cctgcgccgc aagattgccc agcgcatgca ggacgccaaa     4740

cgccgggtcg cgcacttcag ttatgtcgag gaaatcgacg tcaccgccct ggaggccctg     4800

cgccagcaac tcaacagcaa gcacggcgac agccggggca agctgacctt gctgccattc     4860

ctggtacgcg ccctggtcgt ggcgctgcgt gacttcccgc agatcaacgc gacctacgat     4920

gacgaagcgc agatcatcac ccgccatggc gcggtgcatg tgggcattgc cacccaaggt     4980

gacaacggcc tgatggtgcc cgtgctgcgc cacgccgaag cgggcagcct gtgggccaat     5040

gccggcgaga tttcgcgcct ggccaacgct gcacgtaaca acaaggccag ccgtgaagag     5100

ctgtccggct cgaccatcac cctgaccagc cttggcgccc tgggtggcat tgtcagcacg     5160

ccggtggtca acaccccgga agtggcaatc gtcggggtca accgcatggt cgaacggcca     5220

gtggtgatcg acggccagat cgtcgtgcgc aagatgatga acctgtccag ctcgttcgac     5280

caccgcgtgg tcgatggcat ggatgccgcc ctgttcatcc aggccgtacg tggcctgctc     5340

gaacaacccg cctgcctgtt cgtggagtga gcatgcaaca gattatccag actaccctgt     5400

tgatcatcgg cggcggccct ggcggctatg tagcagccat ccgcgccggg caactgggca     5460

ttcctaccgt actggtggaa ggccaggcac tgggcggcac ctgcctgaac atcggctgca     5520

tcccgtccaa ggcgctgatc cacgtggccg agcagtttca ccaggcctcg cgctttaccg     5580

aaccctcgcc gctgggcatc agcgtggctt cgccgcgcct ggacatcggc cagagcgtca     5640

cctggaagga cggcattgtc gaccgcctga ccacaggtgt tgccgccctg ctgaaaaagc     5700

acggggtgaa agtggtgcat ggttgggcca aggtactgga cggcaagcag gtcgaggtcg     5760

atggccagcg tatccagtgc gagcatctgt tgctggcgac cggttccagc agtgtcgaac     5820

tgcctatgct gccgctgggt ggcccggtga tttcctcgac cgaagccctg gcgccgaaaa     5880

ccctgccgca acacctggtg gtggtcggcg gtggctatat cggcctggag ctgggcattg     5940

cctatcgcaa gctgggtgca caggtgagtg tggtggaagc gcgggagcgc atcctgccga     6000

cctacgacag cgaattgacc gccccggtgg ccgagtcgct gaagaaactg ggcatagcgt     6060

tgcacctggg ccacagcgtc gagggctacg aaaatggctg cctgctggcc agcgacggca     6120

agggtgggca actgcgcctt gaggccgacc aggtactggt ggccgtggga cgccggccac     6180

gcaccaaggg cttcaacctg gaatgcctgg acctgaagat gaacggcgcc gccattgcca     6240

tcgacgagcg ctgtcacacc agcatgcaca acgtctgggc catcggcgac gtcgctggcg     6300

aaccgatgct ggcgcaccgg gccatggccc agggcgagat ggtcgccgaa atcatcgccg     6360

gcaaggcccg acgctttgaa ccgacagcga ttgccgccgt gtgctttacc gacccggaag     6420

tggtggtggt cggcaagacc ccggaacaag ccagccagca gggcctggac tgcatcgtcg     6480

cgcagttccc gtttgccgcc aatggccggg ccatgagcct ggaatcgaaa agcggtttcg     6540

tgcgggtggt ggcgcgccgt gacaaccacc tgatcgtggg ttggcaggcg gttggcgtgg     6600

cggtctccga gctatccacc gcgtttgccc aatcgctgga gatgggcgcg tgcctggaag     6660

atgtggccgg taccattcat gcccacccaa cgctcggtga agcggtacag gaagccgcac     6720

tgcgcgccct gggccacgcc ttgcatatct gactcgag                             6758


<210>  16
<211>  26
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  BCKAD.F forward primer for cloning of BCKAD operon

<400>  16
ggcctgtcat gagtgattac gagccg                                            26


<210>  17
<211>  32
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  BCKAD.R reverse primer for cloning of BCKAD operon

<400>  17
cggccctgca ggttcgcggg aatcagatgt gc                                     32


<210>  18
<211>  33
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  AAT.F forward primer for cloning of AAT

<400>  18
aggagatata ccatgaaaag cttttctgta ctc                                    33


<210>  19
<211>  44
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  AAT.R reverse primer for cloning of AAT

<400>  19
agcagccgga tcccctgcag gactagttta ctggctggtg ctac                        44


<210>  20
<211>  39
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  ACX4.F forward primer for cloning of ACX4

<400>  20
caccagccag taagctagca aggagatata ccatggctg                              39


<210>  21
<211>  37
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  ACX4.R reverse primer for cloning of ACX4

<400>  21
tcccctgcag gactagttta caggcgagaa cgggtag                                37


<210>  22
<211>  5652
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  pET16b(Sse) expression vector

<400>  22
ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat       60

aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg      120

tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat      180

gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat      240

tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt      300

aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag      360

cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa      420

agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg      480

ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct      540

tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac      600

tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca      660

caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat      720

accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact      780

attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc      840

ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga      900

taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg      960

taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg     1020

aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca     1080

agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta     1140

ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca     1200

ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg     1260

cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga     1320

tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa     1380

tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc     1440

tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg     1500

tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac     1560

ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct     1620

acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc     1680

ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg     1740

gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg     1800

ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct     1860

ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga     1920

taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg     1980

cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca     2040

tctgtgcggt atttcacacc gcatatatgg tgcactctca gtacaatctg ctctgatgcc     2100

gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc     2160

gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt     2220

acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac     2280

cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga     2340

tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc     2400

ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg     2460

tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca     2520

cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac     2580

tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg     2640

ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga     2700

acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga     2760

agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc     2820

gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg     2880

tcctcaacga caggagcacg atcatgcgca cccgtggcca ggacccaacg ctgcccgaga     2940

tgcgccgcgt gcggctgctg gagatggcgg acgcgatgga tatgttctgc caagggttgg     3000

tttgcgcatt cacagttctc cgcaagaatt gattggctcc aattcttgga gtggtgaatc     3060

cgttagcgag gtgccgccgg cttccattca ggtcgaggtg gcccggctcc atgcaccgcg     3120

acgcaacgcg gggaggcaga caaggtatag ggcggcgcct acaatccatg ccaacccgtt     3180

ccatgtgctc gccgaggcgg cataaatcgc cgtgacgatc agcggtccag tgatcgaagt     3240

taggctggta agagccgcga gcgatccttg aagctgtccc tgatggtcgt catctacctg     3300

cctggacagc atggcctgca acgcgggcat cccgatgccg ccggaagcga gaagaatcat     3360

aatggggaag gccatccagc ctcgcgtcgc gaacgccagc aagacgtagc ccagcgcgtc     3420

ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt     3480

gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat     3540

cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg     3600

tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg     3660

cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg     3720

tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc     3780

gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt     3840

gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc     3900

ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca     3960

ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat     4020

cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc     4080

gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct     4140

cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt     4200

ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac     4260

gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga     4320

ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg     4380

gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag     4440

caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga     4500

gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca     4560

ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg     4620

cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt     4680

gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc     4740

gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga     4800

caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt     4860

gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt     4920

ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg     4980

ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac     5040

agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg     5100

aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca     5160

cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc     5220

ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag     5280

aaataatttt gtttaacttt aagaaggaga tataccatgg cctgcagggg atccggctgc     5340

taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata     5400

accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc     5460

cggatatccc gcaagaggcc cggcagtacc ggcataacca agcctatgcc tacagcatcc     5520

agggtgacgg tgccgaggat gacgatgagc gcattgttag atttcataca cggtgcctga     5580

ctgcgttagc aatttaactg tgataaacta ccgcattaaa gcttatcgat gataagctgt     5640

caaacatgag aa                                                         5652


<210>  23
<211>  1311
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  optimised acyl-coA oxidase 4 (ACX4) from Arabidopsis thaliana for
       expression in pET16b (Sse)

<400>  23
atggctgtcc tgtcaagcgc tgaccgtgcg agcaatgaaa aaaaagtgaa atcttcttac       60

ttcgatctgc cgccgatgga aatgtctgtt gcatttccgc aggcaacgcc ggcatcaacc      120

ttcccgccgt gcacgtcgga ttattaccat tttaacgacc tgctgacccc ggaagaacag      180

gccattcgta aaaaagttcg cgaatgtatg gaaaaagaag tcgcaccgat tatgaccgaa      240

tattgggaaa aagcggaatt tccgttccac attaccccga aactgggtgc gatgggtgtg      300

gccggcggta gtatcaaagg ctacggttgc ccgggtctgt ccattacggc aaatgctatc      360

gcgaccgccg aaattgcacg tgtggatgct tcatgctcga cgttcatcct ggttcatagc      420

tctctgggta tgctgaccat tgcgctgtgt ggctcagaag cccagaaaga aaaatatctg      480

ccgtcgctgg cgcaactgaa cacggtcgca tgttgggctc tgaccgaacc ggataatggc      540

agcgacgcat ctggcctggg taccacggct accaaagtgg aaggcggttg gaaaatcaac      600

ggtcagaaac gttggattgg caatagtacc tttgcggatc tgctgattat cttcgcccgc      660

aacaccacga ccaaccagat caacggtttc atcgtcaaaa aagacgcacc gggcctgaaa      720

gctaccaaaa ttccgaataa aatcggtctg cgcatggtgc agaacggcga tattctgctg      780

caaaatgtgt ttgttccgga tgaagaccgt ctgccgggtg ttaacagttt ccaggacacc      840

agcaaagttc tggcagtcag ccgcgtcatg gtggcttggc aaccgattgg catctctatg      900

ggtatctatg atatgtgcca ccgttacctg aaagaacgta aacagtttgg tgcaccgctg      960

gcagcattcc aactgaacca gcaaaaactg gtccagatgc tgggtaatgt gcaagcaatg     1020

tttctgatgg gctggcgtct gtgtaaactg tatgaaacgg gtcagatgac cccgggtcaa     1080

gcaagcctgg gcaaagcctg gattagttcc aaagcgcgtg aaaccgcaag cctgggtcgt     1140

gaactgctgg gcggtaacgg catcctggcc gattttctgg ttgcaaaagc tttctgcgac     1200

ctggaaccga tctatacgta cgaaggtacc tacgatatta atacgctggt gaccggtcgc     1260

gaagttacgg gcattgcaag cttcaaaccg gctacccgtt ctcgcctgta a              1311


<210>  24
<211>  1368
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  optimised alcohol acyl transferase  (AAT) from Apple for 
       expression in pET16b(Sse)

<400>  24
atgaaaagct tttctgtact ccaagtcaaa cgcctgcaac cagaactgat tacgccagcg       60

aaatcgaccc cgcaggaaac caaattcctg tctgacatcg atgaccaaga gagcttgcgt      120

gtgcagattc cgatcatcat gtgctataaa gacaacccga gcctgaataa gaatcgcaat      180

ccggttaagg ccattcgtga ggccctgtcc cgtgcgctgg tttactatta cccgctggcg      240

ggtcgtctgc gtgagggtcc gaatcgcaaa ctggtggtgg actgcaatgg tgagggtatt      300

ctgtttgttg aggcgagcgc ggacgtcacc ctggaacagc tgggcgacaa gatcctgccg      360

ccgtgtccgc tgttggaaga gtttctgtac aacttcccgg gcagcgatgg tatcatcgat      420

tgcccgctgc tgctgattca agtcacttgt ctgacgtgtg gtggctttat tctggctctg      480

cgcctgaacc acaccatgtg tgatgcagcg ggtttgttgc tgttcctgac cgccatcgca      540

gagatggccc gtggtgccca cgcaccgagc attctgccgg tgtgggaacg tgaactgctg      600

ttcgcacgtg acccgcctcg tattacttgc gcgcaccatg aatacgagga cgttatcggc      660

catagcgacg gcagctacgc gagcagcaac caaagcaata tggtgcagcg tagcttttac      720

ttcggcgcga aagaaatgcg tgttctgcgc aagcagatcc cgcctcacct gatcagcacg      780

tgcagcacct ttgatttgat taccgcatgc ctgtggaagt gccgtacgct ggcgctgaac      840

atcaacccga aagaagccgt ccgtgtgagc tgtatcgtta acgcgcgtgg taaacacaac      900

aatgttcgcc tgccgctggg ctattacggc aatgcgttcg cattcccggc tgctatctct      960

aaggcagagc cgctgtgtaa gaaccctctg ggttacgccc tggagttggt gaagaaggcg     1020

aaagcgacca tgaatgaaga gtatctgcgc agcgtggcgg atctgctggt tttgcgcggt     1080

cgtccgcaat actccagcac gggttcctat ctgattgtga gcgacaatac ccgcgtgggt     1140

tttggtgatg tcaacttcgg ttggggccag ccagtctttg ctggcccggt caaagcattg     1200

gacctgatta gcttctatgt tcaacataag aacaacacgg aagatggtat cttggttccg     1260

atgtgcctgc cgtcctcggc gatggagcgt ttccaacagg agctggagcg cattacccag     1320

gaaccgaaag aggatatttg caacaatctg cgtagcacca gccagtaa                  1368


