. 2022 Feb 17;90(2):e00587-21. doi: 10.1128/iai.00587-21

TABLE 2.

Primers^a

Primer	Sequence	T_a (°C)
Δmcb_VF	GGGGCTTAAAGGGGTAGTGT	49
Δmcb_VR	CCTAACAACGCCACGACTTT	49

ΔmctAB_KF	acacatttcgtacagcctttacactcggtgaattagcggccctagatgcaGTGTAGGCTGGAGCTGCTTC	67
ΔmctAB_KR	ttaaacctcatgttttgtgatatctataatctgtgctttaggtatattatCATATGAATATCCTCCTTAG	67

ΔmctAB_VF	GAAGATATCGCACGCCTCTC	54.5
ΔmctAB_VR	CGCCTGTTTGGCTATATGTG	54.5

ΔcirA_KF	gcagtatttactgaagtgaaagtccgcccggttcgccgggcatcttctcaGTGTAGGCTGGAGCTGCTTC	72
ΔcirA_KR	ctatttcttgtgcatggcctgtgttagcggtcgatgacgatggcgaaacgCATATGAATATCCTCCTTAG	72

ΔcirA_VF	CCCGACGCTTATCGATCAGGG	56
ΔcirA_VR	TGGTCCGGCTTTCTGGGATG	56

cirA_fwd	ggccctttcgtcttcaagaaGTTTCTCCCTTCCTTGCTAAG	57
cirA_rev	taagctgtcaaacatgagaaTCAGAAGCGATAATCCAC	57

pBR322_cirF	TTCTCATGTTTGACAGCTTATC	45
pBR322_cirR	TTCTTGAAGACGAAAGGG	45

pBR322_cirVF	GGGCGACACGGAAATGTTG	53
pBR322_cirVR	GCGCTAGCAGCACGCC	53

Δfur_KF	aaagccaacctgcaggttggcttttctcgttcaggctggcGTGTAGGCTGGAGCTGCTTC	72
Δfur_KR	tctaatgaagtgaaccgcttagtaacaggacagattccgcCATATGAATATCCTCCTTAG	72

Δfur_VF	GCCGCACGTTTGAGGAATTT	52
Δfur_VR	TTTGCCAGGGACTTGTGGTT	52

pBR322_insF	GCAAAAACAGGAAGGCAAAATG	46
pBR322_insR	CTGTCAGACCAAGTTTACTC	46

pBR322_mctA_F	gtatatatgagtaaacttggtctgacagGAACCTACAACACATGTGTAAAACGTCAATG	60

pBR322_mctB_R	tgccttcctgtttttgcTTTTAAACCTCATGTTTTGTG	57

pBR322_mctC_R	tgccttcctgtttttgcGGGGAAGCCCCCTTAGATTAATG	60

pBR322_mctDE_R	tgccttcctgtttttgcATATGCTTGCTTGGGAAATTC	60

pBR322_mctC_F	taaacttggtctgacagATGAATAATCTTATAAAAAAGGAAATCATAGAAAAATTTAAGAAATATAATTTC	53

pBR322_mctE_F	taaacttggtctgacagATATGCTTGCTTGGGAAATTC	58
pBR322_mctE_R	tgccttcctgtttttgcTCACACTACCTTCCTCATATC	58

pBR322_mctD_F	taaacttggtctgacagGTGACTAATTTTAAATCAGACTTAAATC	54
pBR322_mctD_R	tgccttcctgtttttgcCCATTAATCTAAGGGGGC	54

pBR322_insVF	TTTGCAAGCAGCAGATTACG	49
pBR322_insVR	GCCTCGTGATACGCCTATTT	49

ΔmctA_F	CATAAAGCCCGTAATATAC	55
ΔmctA_R	AACACCCCCAATTATATATTTG	55

ΔmctB_F	AAATGAATAATCTTATAAAAAAGGAAATC	57
ΔmctB_R	TTTATAATCCTTAAAGCCCG	57

ΔmctC_F	CCATTAATCTAAGGGGGC	57
ΔmctC_R	TTTTAAACCTCATGTTTTGTG	57

pACYC_PmctA_F	aacgcagtcaggcaccgtgtCGTGAGTTTTCGTTCCAC	64
pACYC_mctA_R	gaggtgccgccggcttccatTTATCTAGAATTACATGAGCAC	64

PmctA_mctB_R	taaaattcaaAACACCCCCAATTATATATTTG	47

PmctA_mctB_F	tgggggtgttTTGAATTTTATTGAAAAGTACATCATC	54
pACYC_mctB_R	gaggtgccgccggcttccatTTAAACCTCATGTTTTGTGATATC	54

PmctA_mctC_R	gattattcatAACACCCCCAATTATATATTTG	47

PmctA_mctC_F	tgggggtgttATGAATAATCTTATAAAAAAGGAAATCATAG	52
pACYC_mctC_R	gaggtgccgccggcttccatTTATATATGTTCAAGTTTTATTTTTTTATAGC	52

pACYC_dTc_F	ATGGAAGCCGGCGGCACC	58
pACYC_dTc_R	ACACGGTGCCTGACTGCG	58

pACYC_VF	GCAAGAGATTACGCGCAGAC	53
pACYC_VR	TAACCAGTAAGGCAACCCCG	53

pRS551_mctA_F	GAATTCTATAACCATTAAAAAACTTGATTACTATCTC	47
pRS551_mctA_R	CCCGGGATCCTTAGAAGAACATCATC	47

pRS551_mctB_F	GAATTCCAAAAGAATCCATATCCAG	47
pRS551_mctB_R	CCCGGGTAAGCAGGATCCTATTTCTCCTATTGAATC	47

pRS551_mctC_F	TAAGCAGAATTCGCTACACAGATTTAAG	49
pRS551_mctC_R	TAAGCAGGATCCATAGTGCAATATATC	49

pRS551_mctE_F	TAAGCAGAATTCGCTGCATAGCTATGCATG	55
pRS551_mctE_R	TAAGCAGGATCCTATGACTGGGATTACTCT	55

pRS551_VF	TGCCAGGAATTGGGGATC	51
pRS551_VR	GTTTTCCCAGTCACGACGTT	51

qPCR_rrsH_F	CGATGCAACGCGAAGAACCT	60
qPCR_rrsH_R	CCGGACCGCTGGCAACAAA	60

qPCR_mctA_3393F (Afwd)	TCTTGGTGCTTACCTCCACCA	60
qPCR_mctA_3556R	TTGCGACAGTTTCATGACCCA	60

qPCR_mctB_4389F (Bfwd)	GCACTTAGCTCCAAATTCGC	60
qPCR_mctB_4567R (Brev)	GCGGAGCTGATACCAAACAG	60

qPCR_mctC_5589F (Cfwd)	TGGCAAATGACAACTTTCCCG	60
qPCR_mctC_5715R (Crev)	GCGCCATCACGTAAGCATTT	60

qPCR_mctD_6365F	AAACTGCATTTTCCATCCACCA	60
qPCR_mctD_6541R (Drev)	CGTCCAGCGAGGATTTTACC	60

Ta, annealing temperature. In primer sequences, lowercase letters indicate homology to the chromosome (for KF/KR knockout primers) or overlapping regions (for Gibson assembly primers). Underlined sequences represent recognition sites for restriction enzymes.