CCCATGGGGCTCTCCCTCAAATGCCAGGGCCGTAACCTAAGTGCACTCGCCTCCCTGCCAACCCTCCCCAGTGGCCATGGCATTGCGGCGAGGAGGACACTCACAACGCCCCGCGGGAGCCCAGCCATCCAAGCCACTTCACCCAGGGCTTCTCATGGATCGTCCGATGAAAAGCACCCACACTCCCTTGCGGCCATGTTGCCCCCTGGCCTCTTGAGTTCAAGGTTTTGCGCGACTAGCAACCTGGGGACAGCC

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CCCATGGGGCTCTCCCTCAAATGCCAGGGCCGTAACCTAAGTGCACTCGC CTCCCTGCCAACCCTCCCCAGTGGCCATGGCATTGCGGCGAGGAGGACAC TCACAACGCCCCGCGGGAGCCCAGCCATCCAAGCCACTTCACCCAGGGCT TCTCATGGATCGTCCGATGAAAAGCACCCACACTCCCTTGCGGCCATGTT GCCCCCTGGCCTCTTGAGTTCAAGGTTTTGCGCGACTAGCAACCTGGGGA CAGCCTGCTCCCCTCCCTTCCAGGAGTGGCGATGGAGCTATTTCCCATCA M E L F P I Q AATCCTGTTCCTTGTGGTGTTGATGAACCGGTACATCCTGGGGCCGCTCA I L F L V V L M N R Y I L G P L M TGCGGCGGGTCCGAGGCGATCAGTTCGATGCGACGAATGATGCCTACGAA R R V R G D Q F D A T N D A Y E CCCACGGTGGCCATCATCATCCCGCTCTTCAACGAGGGCGAAGGCATCTA P T V A I I I P L F N E G E G I Y TCACGCCATCCGCAGCCTGCTGTTGCAAGATTACCCGCCTGACAAATTGT H A I R S L L L Q D Y P P D K L S CGATCGTTGTCGTCGATGATTGCTCCAAGGACGACAGCTATGTCTGGGCA I V V V D D C S K D D S Y V W A CTGAAGGCCGCCGAGCAGCATCCCAACGTCATGGTGATGCGCAACCCGGA L K A A E Q H P N V M V M R N P E GAACATGGGCAAGCGCAAGGGCATCAACCGGGGCGTTCGCGCCACCCAGT N M G K R K G I N R G V R A T Q S CGGAGATCATCGTTTCGGTGGACTCGGATGTCATCGTGGACCGCGGCGCC E I I V S V D S D V I V D R G A GTACGGCAGCTCGTCCGCCGGTTCCTCCACCCGCGCATCGCCGCGGTGGG V R Q L V R R F L H P R I A A V G CGGAAGAACCTACGTGACCAACCGTCATCAGAACTGGATGACGCGGATGA G R T Y V T N R H Q N W M T R M I TCGAGATCAAGTTTCACTTCGCCCAGGAGTGGCTCAAGGATCTGGAGCGG E I K F H F A Q E W L K D L E R GGTTTCCGCTCGGTGATGTGCCTGTCGGGGTGTCTGACAGCGTACCGCCG G F R S V M C L S G C L T A Y R R CCACGTGCTGGAGGAGCTGGAACCCATCCTGGAAGCGCGCTCCATCGCCG H V L E E L E P I L E A R S I A G GGGTCGCCATCAAGTATGGCGAGGACCGCTTCCTCACCCGCCAGATCATC V A I K Y G E D R F L T R Q I I AAGGCGGGCTACGAGACCATCTACACCACCGCCGCGGTCTGCTTCACCGC K A G Y E T I Y T T A A V C F T A CACCCCGGCCAACATCGCGGGTTACTTCGCACAGCAGCTCCGCTGGCGGC T P A N I A G Y F A Q Q L R W R R GCTCCAACCTGGTGGACATGTTGGGGGGACTCAGCCACGCCTGGCGGCTG S N L V D M L G G L S H A W R L CACCCCGTGGTCGCGGTCCACTATGTTTCCCAGTTTGGGTTGCTGCTCTC H P V V A V H Y V S Q F G L L L S CTACCCGGTTGTCATCGTCCACAACATCCTGACTGGTGAGTTCTGGGACA Y P V V I V H N I L T G E F W D I TCCTGGCCATGCACGTGCTCACGGTCGGGATCATGGGGTTCATCTACCGC L A M H V L T V G I M G F I Y R TGGGAAACACGCCACCTGCCTGATGACCAGCGTGTCCCAGGCCTGAGCTT W E T R H L P D D Q R V P G L S F TCTTCCCATGGCCTTGATGATGCCCATCACCTACGCGCTCTTCACGCCCC L P M A L M M P I T Y A L F T P L

50 100 150 200 250 300 350 400 450 500 550 600 650 700 750 800 850 900 950 1000 1050 1100 1150 1200 1250 1300 1350 1400 fbfA

pSM24

225

(101)

(102)

(103)

(104)

ACGCCCTTCGCGCGCCCCTCGACTAGCCCCGATAGAAGCCCGCCTGCCCA TCACGACGCGCCGAAAGCGCTGTACCTCCTGAGGAGACCGACCCACCCAG CTGCCACTGACGCCCACTCTGAGTCCATCGCCTTCCCGAGAGCGGCAGGC GGTGCCGCGCCGTCCGGCCCGTCAGCCTGGTAGCCAGTAGCCGCCGTACC CACGCCACGTCCCGCCGGCGTTCGGCGAGGTCCCGGTAGTCCCTCCGCAG GTGCTCAACCCTGCGGTCGTGGAACGCGAGTGCAGGCCGCGGCCTGAACT CCTGGGGGTAGACGACTGCGTGGTGCCGTTCGCCTACCCCGAGACAGTGC CCGAACACGGTCAGTTGACGCGGTCAGGCACGGTCAAACCGCCTGAGGCG CAGATTCGGGAGTCTAAAACAACCGTTGACCGCGCCTAACCGCATCTTCC TCGGGTCCTCATGCCCAAGTTCAGGGCATGAGGAGCGGTTATCTCCCCGG ACCTTAGCTCTCCCTTTGGAGCTTCTAAGGTCCGGGAAGCAGAAAAGTCG AGGGCCCGAAGAGGGAGCCGCACTAGAGGCAGAGGTCCTACAGAGGTCTT TAGACCCTGCGGCGGAGTAGCTCGTCCGACTACCGGCGCAGGAAGCGACG GCCCGAGTCCACGTACATCGCGACCCACCACCTCTCCAAGAGCACGGGCC GGTCAAGGACCTACCGGAATCGCCCGCGGGTAGCGTAGCGGTCGCCGCTC GTCTTCCACACGGCCTCCTGCACCTCGCGAGCACAGTGGCCGTCGGGGCG GGCGAAGACGCGGAAGCGGTAGGTCGCCCTCTGTCGGCGGGCCCAGCGAC CAACCGGCAACAGGGCCATCTCCTGTGCGCCTGGGAAGTCTACGGCTAAC CAGCGCTCGTTGCGCCGGTCGGCGAAGCAAGTACCCGTGGGCAGCTAGTG CGGGTGGGAACCCGTGCCACTGCACGGGGACGGTGAGACTAACGGCCTAC ACCTCGACAGGAGCGGCCTTCAGGTGCAGGCAGGTGAGGTTGCGCTCCTA GAGGGGTGACGCGTCCGGGCGCAGAGGTGGCTCGTCGTGGTACCGGACCT ACGCGGTCAGGTCGAACGACCGGCGGAGCTGGTCGGCGAGCATGAGGAGC TTCAGCATCTTGCTGTACCGCCCGAAACCTCGGAACTCGTCGAACTACCA CTGGCCGTCGAACAGCTCCTGCGGGGTGAGGTGGCGGTGGAAGTCGTCGA ACAACTTCTGCCACTCCTGCAACAAGTGCCAGAAGCACCTGTCTAGGAAG TCCAGGCCGGAACTCCGCGACCTACAGGAGCACGCAGTCGCACAGGTCGG CGGCGACTGGCTTGGCCCCGTGGTCCATCTAGGCGTCGTGCCTGAGGAAC

50 100 150 200 250 300 350 400 450 500 550 600 650 700 750 800 850 900 950 1000 1050 1100 1150 1200 1250 1300 1350 1400 attB

pSM49

ire

230

(105)

GACTACTACCACGACCCGAAGACGGCTAACCGGGCCCGCATCGGGAGCTA CTTGGACCCTTGCTTGAGGAGGTTGTAACCCTGGCGGAAGGCAAAGTGGA GGAGGAACCGGACGAACGGCGCACGCTCGTTGAGCCTGACGGCGACAAGC GGGGTGCGGAAGCGCGGGCTGAAGCTCGACTGGCCTCGGAAGTGCGCGAG CGCCATGTCGAGCGGTAGCCCGGTGAACTTGTACTACAGGTGGAGGGTAG GCGAGAACGGTGCGAACTACATGCCGGACTGGAAGTGCGAGTAGTGGCTC GGGAGGAGTCGCTGCCGCGTGCGGGAAGCTCTTATTTGCCGTCTTCGAGC GAGGGGGCCTTGGCGTCACTCGCCGGGTTCGCCTTGCGCGAACCTTCGAC CGGCGCGCTGTGGAAGCGCATGTGCCGGAACCCCGACGTGGACTCCTCCG CCCGGTGGAGGAGCCGCCACATCTGCCGGAACAGCTCGCCACCAACAGGG GAGGCTCCCCGACGACGACCCTCTTGGACGGACGTGGGCGAGCGCCTCGC GACCAAGGGGCACTTCGCGGCGGTCAAGGAGCGCGTGGTCCCGCGCCTAC TCCAGTGCGTCAGGCCGGAGATGCGACTCTGTGCGACTGGAGGGTGGCAG TGACGACCTTACTTAGGGCAGGACGTAGAGTTGGAGCACTTCTGCTCCGA GTACCACCTGCACGGCGTCCTGCTTCCCGTCGCCCTATCAAGGCGGAGAC GGCTGGATGAAGAATAGGGGGAACCGTAGGACCCTTAAAAGTCATCCACA GCTGAAGCAAAAGCCACAGCGCCATCGACCGCCGAACTCGCGCGGTATCC CACAAGGGCGCAAGCCGAGGCCGCGTACCCCGAGCCACAGGTACGCTGCG TTCTACCCGTCGTGCAGGCGAGCCCGCAAGAAGTAGTAGAGGAAGGTCGG GAGGGCAGCGAAGAACAACGAGCACGCTAGGAAAAGCTTGAGGAGTAGAC CGCTCGCTTAGGGCTCGTCCTCGCGTTGTCGGAGGTACAGGACTCCGTTC TCGAAGCCGGGGTGCAAGAGCTGGAGGTGGCGCATGTAGCGCTTCCGCGT CCATTCGGAGTTGAGCGAGACGGTCTCAGCGCACGCGAGGAAGTGGGTCC CCGCGAAGAGCATTTCGCCGCGCACCAGTGGACTTCAACCCGCGGTCGGA CTATCTCGGAGACCATCCGCGACTCCTGAAGGTCCGGCAGGCACTGCTAT GGGTACTTCCCCAAGTCGAACCTCCACTTGCCTGGCCCTCGCGGTCCGCC GAGCCCGTAGAAGCCGCCCCTGAAGTCTATCGACGGGCTGTTGTTGGTGA GGACTAACGCCACATGGGTGGACTTACGGACCCGCTGCCGTATAGGGTCC

1700 1750 1800 1850 1900 1950 2000 2050 2100 2150 2200 2250 2300 2350 2400 2450 2500 2550 2600 2650 2700 2750 1450 1500 1550 1600 1650

2800 231

(106)

GGTCGCGAGCGCGTCTACTACAGCTAGACGACGACTCATGCGGCGGACTC GTCCACGAAGTTCCGCCGACCGCCCAGGAACTTGGCTCCGAACCCGAAGA AGAGCCCGAGTCCCGGACAGCCAACTGCCCCGCAGGGGTAGAACGCACGC GGAGGAGACGCGGGACCGACTCGCGCTGCCAGAGAAGGTTCTCGTAGGAG AGCTTCTTGCATCGACCCCCTAAGGCGACCTCCGCCGGTTTATAAGGGCC CTATTCAGGCCGGCTCTTCGCGTCGTAGATAGGCAGAGAGAGCCAAAGTC TTCGTGTACCAACGCCCAGTGGAGCAGCCACTGGCACGCGGGCCTCGGCG GAATGTCTGGCCATCTGCCAGGAACGGGCTACGTCGTGCTCCTCCAACTG GCTACTTGTGGCTAGGGTCCTACTGCCAGGTTCTCTTGAGAGGACCTGTG CCGGAGCGGAGGGGTCTCGGGGGAAGGACGGGACCTCGGGAACGGGTGGC CGTCGCACATGGCTGCAGACAGGCA

2850 2900 2950 3000 3050 3100 3150 3200 3250 3300 3325

232

(107)

CAATCGGCATCTGAAGGGTCCGCGTGTCCTCTACCGGGACAACGGCCAAC CAGCGACCCGGGCGGCTGTCTCCCGCTGGATGGCGAAGGCGCAGAAGCGG GCGGGGCTGCCGGTGACACGAGCGCTCCACGTCCTCCGGCACACCTTCGC TCGCCGCTGGCGATGCGAGGGCGCCCGCTAAGGCCATCCAGGAACTGGCC GGGCACGAGAACCTCTCCACCACCCAGCGCTACATGCACCTGAGCCCGGC AGCGAAGGACGCGGCCATCAGCCTGCTCGATGAGGCGGCGTCCCAGATTT CTGGAGACATCCTGGAGACGGAGATCACGCCGAGGGAGAAGCCCGGGAGC TGAAAAGACGAAGGGCCTGGAATCTTCGAGGTTTCCCTCTCGATTCCAGG CCCCTCTATTGGCGAGGAGTACGGGACTTGAACCCGTGGCCTCCGGCGTG ACAGGCCGGCGTTCTAACCAACTGAACTAACTCCCCACAACTTTGGGCGG AACAGGGATCGAACCTGTGACCCTCGCCTTGTAAGGGCGACGCTCTACCG CTGAGCTATCCGCCCTTGGCAACCGCCGCGTCGATGGGCGGCCTTGTATC GACCACCTCCCCCTGTGTCAAGCATACGTTTCTGCCGGTTTGATCATTCC CTGCGCGCTGCGTCGCGTCGAGCCAGGCGCTGTCCACTGCCGGTCCAGCG CCCTGCGTCAATCGGCCCCTCCTGGCGGGGGCGCGTTGACTCGGCCGCTC AAGCAGTGGTGGAAGGTTGCAGTACGGGCGAGCGCGGGCCCCCTGTAGCG CTCTTCGCAACCGCGCTGGGAGTTGTACCCATCATGATGACGTCCATCAG AAACCTTGGCAGGGCGTGGAGCGCCCTGCCGTGGCGGCCCCTGCTTGCGG TCGGCCTC

trnD1

trnV1

50 100 150 200 250 300 350 400 450 500 550 600 650 700 750 800 850 900 908 attB

pSM54

233

(108)

f1

lac New

col Amp

Untitled 1 10.00 kb Amp^r

ColE1 origin

MCS LacZ

ca 2.9 kbp

pBS SK (+/-) f1(+) origin

Sac I Kpn I

f1(-) origin pBS SK (+/-) /

Sac I Kpn I

pBS KS (+/-)

234

(109)

New MCS

rep amp Untitled 1

10.00 kb Amp^r

MCS LacZ

ca 2.6 kbp

pUC18

trp-lacZ Kan^r

Sfi I Eco RI Sma I Bam HI Cla I Hind III Not I Xho I

Sfi I Sma ICla I Hind III Not I

mini-Tn5-lacZ1

rep(pMB1) pUC18

cm

ori

tc mob

Untitled 1 10.00 kb

Tet^r pSUP102

ca. 6 kbp

mob pSUP102

ORI Cam^r

Eco RI

Xba I Cla I

Eco RV Bam HI Sal I

235

(110)

pUC4KIXX

4 kbp Kan^r lacZ lacZ

ori

Kan^r Ble^r

Eco RI Bam HI Sac I Sma I Hind III Xho I Eco RIBam HISac ISma IHind III

Xho I

Sma I Sal I

Sph I

1.5 kbp

Amp^r

amp

ori New ap

Untitled 1 10.00 kb pBR322

ca 4.3 kbp

Amp^r

Tet^r

ORI ROP

pBR322 pUC4KIXX

236

(111)

pBIM9

ca. 8.4 kbp

Mx8 ColE1

f(-)

Xho I Sac

I

intP attP

Sma I

Amp^r

pLJS49 22.4 kbp

Mx8

P1 Amp

Kan ColE1

Eco RI Bgl II

Sac I

Xho I

intP attP

Sma I

2.9 kbp

r

pLJS49 pBIM9

237