PRIMER NAJAD

Syarat mengunakan primer dari jurnal

1. Cek dengan DNA calculator
2. GC conten 40-60
3. Primer 18-20
4. Apakah spesific band? Spesifik dengan phasil fast PCR 507 bp

Pseudomonas fluorescens ice nucleation gene (inaW)

GenBank: X04501.1
GenBank Graphics
>gi|45507|emb|X04501.1| Pseudomonas fluorescens ice nucleation gene (inaW)
ATGAATGCATAATGGGCACTGGATGACATAGATGGCGTACGGTTGCCATTTATGGGCCCCAACGTGACCG
ACCATCGCTTGGGTATTCTTTAATTCCCCAGCGGTAATCTGGAAGGAAAACTACTTCCGCCAACTCATCC
TTCTGTCAGTGCCACGTCCGACAGCAATGACCTCGATCCCCTCTCGCGACGTCTCGCCTTTCTACTACCC
CTTGCTTAAGCAGATTTATGCATAAAAGAGTATGGCGTTGACTCATCATTTTACGAGACAAGCCCAGTCA
GAATTTATCGGCATACTGCGGACCGTCTACCCCCCAATGACCGTTGACCTCATCCGGCTTCTCATTACCT
ATTTTGCATCCGGCGACTGGATGGTGCTAACAGATCATCGGCGTGCGCACCTGGCTGACACAGATTCACC
AACANACCTTAGCTTTGCTGTAGTTGGAAATCAGGGCAAAATATTCTGACAAATGCGCCAACCGTAGCTT
GGTTGAGTTACATGCCAGCAACTTCATCTGATGAGTATTGTTCTGCCCTCAAGATCGACGTATTTTTTGC
AGTATCGCGCCATAATATTTACTCGTTAGTTTGAGCATTAATGGCTGCATAGACATCGTATGTGAAAATG
TATTATGGACGATCCCCGTTAAAGGACTTAATAGAAATGAGGCATAAGGGTAATTCAAAATGAAAAGTGA
AAAAGTTCTGGTTTTACGGACATGCGCTAATAATATGACCGACCATTGCGGTCTGGTATGGCCTATTTTA
GGGCTTGTCGAATGCAAGTTTTGGGAGCCCACAATAAAACTCGAAAATGGGTTGACCGGCGCACTGTGGG
GACAAGGCTCAAGTGCCCAGTTGAGCATGAATGCAGACGCAAAATGGGTTGTTTGTGAAGTGACAATGGG
CGACCTGATTTTCTTGGAAAATAATGAGGGGGTCAAGTTTCCTCGTGCAGAAGTAGTTCATGTCGGAACA
CGAAGTAGCGCGCTAGGCTACATTTCTGACAATGTTTCCAAGCATGAAGCATGTTCAAGTAATCTGATTG
AGAAATTTACTTTTTCTGATGTTAAATCAGAGACGAGGAATATTTCTCCCGCGCTGCCGGTTACTGTTGA
CAACATGCCTAATGGCGTCAATCGCAGCACAACTGTACGCAATACGCAAACGCTGGAAACGGCCGTTTAT
GGCAGCACGCTCACTGGTGCTAATCAAAGCCAGCTCATTGCAGGCTACGGTAGCACCGAAACCGCTGGAG
ATAGCAGCACTCTTATTGCGGGATACGGCAGTACCGGAACTTCGGGCTCTGATAGTTCGATCATTGCGGG
TTACGGCAGCACAGGCACCGCCGGCTCCGATAGCTCACTCATTGCCGGATATGGCAGTACGCAGACTGCA
GGCGGAGATAGCTCCCTAACTGCCGGTTACGGTAGTACCCAAACGGCTCAGGTGGGCAGTAATCTCACCG
CTGGCTATGGCAGCACCGGCACTGCGGGTCCTGACAGCTCACTTATCGCAGGTTATGGCAGCACGCAGAC
TGCTGGGGGCGAAAGCTCCCTGACCGCCGGTTACGGCAGTACCCAGACGGCTCAGGTGGGCAGTGATCTA
ACCGCTGGCTATGGCAGCACCGGCACTGCAGGTTCCGATAGCTCGCTCATTGCCGGATATGGCAGTACGC
AGACTGCAGGCGGAGATAGCTCCCTAACTGCCGGTTACGGTAGTACCCAAACGGCTCAGGTGGGCAGTAA
TCTCACCGCTGGCTATGGCAGCACCGGCACTGCGGGTCCTGACAGCTCACTTATCGCAGGTTATGGCAGC
ACGCAGACTGCTGGGGGCGAAAGCTCCCTGACCGCCGGTTACGGCAGTACCCAGACGGCTCAGGTGGGCA
GTGATCTAACCGCTGGCTATGGCAGCACCGGCACTGCAGGTTCCGATAGCTCGCTCATTGCCGGATATGG
CAGCACGCAGACTGCTGGGGGCGAAAGCTCCCTGACCGCCGGTTACGGCAGTACCCAGACAGCTCAGGTG
GGCAGTGATCTAACCGCTGGCTATGGCAGCACCGGCACTGCAGGTTCCGATAGCTCGCTCATTGCCGGAT
ATGGCAGCACGCAGACTGCAGGTGGAGATAGCTCTCTGACTGCCGGTTACGGTAGTACCCAAACGGCTCA
GGTGGGCAGTGATCTCACCGCTGGCTATGGCAGCACCGGCACTGCAGGTTCCGATAGCTCGCTCATTGCC
GGATATGGCAGCACGCAGACTGCAGGCGGAGATAGCTCTCTGACTGCCGGTTACGGCAGTACCCAAACGG
CTCAGGTGGGCAGTGATCTCACCGCTGGCTATGGCAGCACCGGCACTGCAGGTTCCGATAGCTCGCTCAT
TGCCGGATATGGCAGTACGCAGACTGCAGGCGGAGATAGCTCCCTAACTGCCGGTTACGGCAGTACTCAA
ACGGCTCAGATGGGCAGTAATCTCACCGCTGGCTATGGCAGCACCGGTACTGCAGGTTCCGACAGCTCGC
TCATTGCCGGATATGGCAGCACGCAGACGGCAGGCGGAGATAGCTCCCTGACCGCTGGTTACGGCAGTAC
TCAAACTGCCGGTCACGGAAGCATCCTGACTGCCGGATACGGCAGTACGCAGACGGCACAGGAAGGAAGT
TCACTCACCGCAGGATATGGCAGCACGAGCACAGCCGGTCCTGAGAGTTCGCTGATCGCTGGCTACGGCA

GCACCCAGACTGCAGGACACGAGAGTACACTCACTGCTGGCTACGGCAGCACCCAAACAGCTCAGGAGGA
TAGTTCACTTACTGCGGGATATGGCAGTACATCAACCGCAGGATTCAACAGCTCGTTAATCGCAGGCTAC
GGTAGTACTCAAACCTCTGGGTATGAGAGCATCCTGACCGCCGGCTACGGCAGTACGCAAACGGCCCAGG
ATAATAGCTCTCTGACCACCGGCTACGGCAGTACATCGACTGCGGGTTATCAAAGCTCGTTAATTGCGGG
TTACGGCAGCACCCAGACAGCAGGATATGAATCTACGCTAACTGCAGGTTACGGCAGTTGCCAGACGGCT
CAAGAGCAAAGTTGGCTAACTACCGGTTACGGGAGCACATCAACTGCTGGCTACGAAAGCAGGCTGATTG
CTGGATATGGCAGTACTCAAACCGCAGGCTACAAAAGCATTCTAACCGCAGGCTACGGGAGCACTCAGAC
CGCCCAGGAAGAGAGTTCGCTTACGGCCGGTTATGGCAGCACTTCGACTGCGGGCTATGCAAGCTCCCTC
ATCGCTGGTTATGGAAGCACCCAGACCGCAGGTTACGACAGTATCCTGACCGCAGGTTATGGCAGCACGC
TAACCGCTCTGGATAGCAGTACGCTAACCGCAGGCTACGGTAGTACGGAAACAGCAGGATTTGGCAGTTC
GTTGATGGCCGGTTACGGCAGTTCGCAGATCGCCGGTTATGGGAGCACGTTAACTGCGGGCTATGGTAGT
ACCCAGATGGCAGAGCGGGATAGCACTCTCACTGCTGGGTATGGCAGCACCGGCACTGCGGGGCAGGACA
GTTCCCTGATTGCCGGTTATGGCAGCAGCTTGACCAGCGGCATGCGCAGCTATTTGACAGCCGGTTACGG
CAGCACTTTAATCAGCGGACTTCAGAGTGTATTAACGGCGGGGTACGGTAGCAGCCTTACTTCAGGCATT
CGCAGTAGCCTGACTGCGGGATACGGCAGCAACCAGATTGCGAGTCATAAGAGTTCTCTTATTGCGGGCC
ATGAAAGCACTCAGATTGCAGGGCACAAAAGTATGTTGATTGCCGGCAAGGGCAGTTCGCAAACAGCTGG
TTCTCGCAGCACGCTGATAGCTGGCGCTAATAGCGTCCAGATGGCAGGGGATCGTAGCAGGCTTACTGCC
GGTGCAAACAGCATCCAAACAGCGGGAGACCGCAGCAAGCTCCTGGCAGGCAGCAACAGTTATCTGACTG
CTGGCGACCGGAGCAAACTCACCGCCGGCGACGATTGTGTTCTGATGGCAGGGGATCGCAGCAAGCTGAC
GGCGGGTAAGAACTGCGTATTGACCGCCGGCGCTGACAGCAGACTCATAGGGAGCCTCGGCTCAACGCTC
TCAGGTGGGGAGAACTCTACCCTGATTTTCCGATGCTGGGACGGCAAGCGTTACACCAATGTCGTCGTCA
AGACCGGTACGGACGAGGTAGAGGCAGACGTTCCGTATCAAATTGACGAAGACAGTAATGTTCTAATTAA
GGCAGAGGACAATAGCGATACTCCGGTGGACCAGTCTCAGATTCAGCCATGAGACTTTTCTTTAAGCGAA
GGAGGAGCCTAGGCTGAGCTTGCAGAGATGCTCTGCGGGTAAGGTGGCCGGGGTGGATGTTGACGCATCC
TGACCGGCCGCTCCTTTTGGTTTCGAGGTAAGACTTGTTTTGCGCAGGCACGGCGATCCCGCAAGGCATT
CATGCCTTAGTCGGCGTAACCATTAATTGAGTTGAATCGTAGAGTGTGCAGAGTTCAGAAGGCCCGTGTT
CCGCAAGCCAGGCCGATGACGCAGAGAATGCTTAAGGCAAGCGAGTGGAATGCTGAAGAGGCGCTGACGC
GCCCCGCTAATCAGCACCATTGAAACGTATTCAGATC

FASTPCR
>atgaatgcataatgggcactggatgacatagatggcgtacggttgccatttatgggccccaacgtgaccg
accatcgcttgggtattctttaattccccagcggtaatctggaaggaaaactacttccgccaactcatccttctgtcagtgcc
acgtccgacagcaatgacctcgatcccctctcgcgacgtctcgcctttctactaccccttgcttaagcagatttatgcataaa
agagtatggcgttgactcatcattttacgagacaagcccagtcagaatttatcggcatactgcggaccgtctaccccccaatg
accgttgacctcatccggcttctcattacctattttgcatccggcgactggatggtgctaacagatcatcggcgtgcgcacct
ggctgacacagattcaccaacanaccttagctttgctgtagttggaaatcagggcaaaatattctgacaaatgcgccaaccgt
agcttggttgagttacatgccagcaacttcatctgatgagtattgttctgccctcaagatcgacgtattttttgcagtatcgc
gccataatatttactcgttagtttgagcattaatggctgcatagacatcgtatgtgaaaatgtattatggacgatccccgtta
aaggacttaatagaaatgaggcataagggtaattcaaaatgaaaagtgaaaaagttctggttttacggacatgcgctaataat
atgaccgaccattgcggtctggtatggcctattttagggcttgtcgaatgcaagttttgggagcccacaataaaactcgaaaa
tgggttgaccggcgcactgtggggacaaggctcaagtgcccagttgagcatgaatgcagacgcaaaatgggttgtttgtgaag
tgacaatgggcgacctgattttcttggaaaataatgagggggtcaagtttcctcgtgcagaagtagttcatgtcggaacacga
agtagcgcgctaggctacatttctgacaatgtttccaagcatgaagcatgttcaagtaatctgattgagaaatttactttttc
tgatgttaaatcagagacgaggaatatttctcccgcgctgccggttactgttgacaacatgcctaatggcgtcaatcgcagca
caactgtacgcaatacgcaaacgctggaaacggccgtttatggcagcacgctcactggtgctaatcaaagccagctcattgca
ggctacggtagcaccgaaaccgctggagatagcagcactcttattgcgggatacggcagtaccggaacttcgggctctgatag
ttcgatcattgcgggttacggcagcacaggcaccgccggctccgatagctcactcattgccggatatggcagtacgcagactg
caggcggagatagctccctaactgccggttacggtagtacccaaacggctcaggtgggcagtaatctcaccgctggctatggc
agcaccggcactgcgggtcctgacagctcacttatcgcaggttatggcagcacgcagactgctgggggcgaaagctccctgac
cgccggttacggcagtacccagacggctcaggtgggcagtgatctaaccgctggctatggcagcaccggcactgcaggttccg
atagctcgctcattgccggatatggcagtacgcagactgcaggcggagatagctccctaactgccggttacggtagtacccaa
acggctcaggtgggcagtaatctcaccgctggctatggcagcaccggcactgcgggtcctgacagctcacttatcgcaggtta
tggcagcacgcagactgctgggggcgaaagctccctgaccgccggttacggcagtacccagacggctcaggtgggcagtgatc
taaccgctggctatggcagcaccggcactgcaggttccgatagctcgctcattgccggatatggcagcacgcagactgctggg
ggcgaaagctccctgaccgccggttacggcagtacccagacagctcaggtgggcagtgatctaaccgctggctatggcagcac
cggcactgcaggttccgatagctcgctcattgccggatatggcagcacgcagactgcaggtggagatagctctctgactgccg
gttacggtagtacccaaacggctcaggtgggcagtgatctcaccgctggctatggcagcaccggcactgcaggttccgatagc
tcgctcattgccggatatggcagcacgcagactgcaggcggagatagctctctgactgccggttacggcagtacccaaacggc

tcaggtgggcagtgatctcaccgctggctatggcagcaccggcactgcaggttccgatagctcgctcattgccggatatggca
gtacgcagactgcaggcggagatagctccctaactgccggttacggcagtactcaaacggctcagatgggcagtaatctcacc
gctggctatggcagcaccggtactgcaggttccgacagctcgctcattgccggatatggcagcacgcagacggcaggcggaga
tagctccctgaccgctggttacggcagtactcaaactgccggtcacggaagcatcctgactgccggatacgGCAGTACGCAGA
CGGCACAGgaaggaagttcactcaccgcaggatatggcagcacgagcacagccggtcctgagagttcgctgatcgctggctac
ggcagcacccagactgcaggacacgagagtacactcactgctggctacggcagcacccaaacagctcaggaggatagttcact
tactgcgggatatggcagtacatcaaccgcaggattcaacagctcgttaatcgcaggctacggtagtactcaaacctctgggt
atgagagcatcctgaccgccggctacggcagtacgcaaacggcccaggataatagctctctgaccaccggctacggcagtaca
tcgactgcgggttatcaaagctcgttaattgcgggttacggcagcacccagacagcaggatatgaatctacgctaactgcagg
ttacggcagttgccagacggctcaagagcaaagttggctaactaccggttacgggagCACATCAACTGCTGGCTACGAAAgca
ggctgattgctggatatggcagtactcaaaccgcaggctacaaaagcattctaaccgcaggctacgggagcactcagaccgcc
caggaagagagttcgcttacggccggttatggcagcacttcgactgcgggctatgcaagctccctcatcgctggttatggaag
cacccagaccgcaggttacgacagtatcctgaccgcaggttatggcagcacgctaaccgctctggatagcagtacgctaaccg
caggctacggtagtacggaaacagcaggatttggcagttcgttgatggccggttacggcagttcgcagatcgccggttatggg
agcacgttaactgcgggctatggtagtacccagatggcagagcgggatagcactctcactgctgggtatggcagcaccggcac
tgcggggcaggacagttccctgattgccggttatggcagcagcttgaccagcggcatgcgcagctatttgacagccggttacg
gcagcactttaatcagcggacttcagagtgtattaacggcggggtacggtagcagccttacttcaggcattcgcagtagcctg
actgcgggatacggcagcaaccagattgcgagtcataagagttctcttattgcgggccatgaaagcactcagattgcagggca
caaaagtatgttgattgccggcaagggcagttcgcaaacagctggttctcgcagcacgctgatagctggcgctaatagcgtcc
agatggcaggggatcgtagcaggcttactgccggtgcaaacagcatccaaacagcgggagaccgcagcaagctcctggcaggc
agcaacagttatctgactgctggcgaccggagcaaactcaccgccggcgacgattgtgttctgatggcaggggatcgcagcaa
gctgacggcgggtaagaactgcgtattgaccgccggcgctgacagcagactcatagggagcctcggctcaacgctctcaggtg
gggagaactctaccctgattttccgatgctgggacggcaagcgttacaccaatgtcgtcgtcaagaccggtacggacgaggta
gaggcagacgttccgtatcaaattgacgaagacagtaatgttctaattaaggcagaggacaatagcgatactccggtggacca
gtctcagattcagccatgagacttttctttaagcgaaggaggagcctaggctgagcttgcagagatgctctgcgggtaaggtg
gccggggtggatgttgacgcatcctgaccggccgctccttttggtttcgaggtaagacttgttttgcgcaggcacggcgatcc
cgcaaggcattcatgccttagtcggcgtaaccattaattgagttgaatcgtagagtgtgcagagttcagaaggcccgtgttcc
gcaagccaggccgatgacgcagagaatgcttaaggcaagcgagtggaatgctgaagaggcgctgacgcgccccgctaatcagc
accattgaaacgtattcagatc

>PRIMER FORWARD
GCAGTACGCAGACGGCACAG
>PRIMER REVERSE
TTTCGTAGCCAGCAGTTGATGTG

____________________
In silico Primer(s) search for:
atgaatgcataatgggcactggatgacatagatggcgtacggttgccatttatgggccccaacgtgaccg
primer forward 5'-gcagtacgcagacggcacag
Position: 2562->2581

100%

Tm = 61,0C

5-gcagtacgcagacggcacag->
||||||||||||||||||||
cggcagtacgcagacggcacaggaag
primer reverse

5'-tttcgtagccagcagttgatgtg

Position: 3046<-3068

100%

<-gtgtagttgacgaccgatgcttt-5
|||||||||||||||||||||||
agcacatcaactgctggctacgaaagcag
>primer forward 2562->2581
5'-gcagtacgcagacggcacag
>primer reverse 3046<-3068
5'-tttcgtagccagcagttgatgtg
PCR product size: 507bp Ta=64C

Tm = 57,2C