Utilizing Hind III and EcoR V Restriction Enzyme with Pet41 and the following gene of interest... a tgaaacaaca aaaacggctt tacgcccgat tgctgacgct gttatttgcg 61 ctcatcttct tgctgcctca ttctgcagca gcggcggcaa atcttaatgg gacgctgatg 121 cagtattttg aatggtacat gcccaatgac ggccaacatt ggaagcgttt gcaaaacgac 181 tcggcatatt tggctgaaca cggtattact gccgtctgga ttcccccggc atataaggga 241 acgagccaag cggatgtggg ctacggtgct tacgaccttt atgatttagg ggagtttcat 301 caaaaaggga cggttcggac aaagtacggc acaaaaggag agctgcaatc tgcgatcaaa 361 agtcttcatt cccgcgacat taacgtttac ggggatgtgg tcatcaacca caaaggcggc 421 gctgatgcga ccgaagatgt aaccgcggtt gaagtcgatc ccgctgaccg caaccgcgta 481 atttcaggag aacacctaat taaagcctgg acacattttc attttccggg gcgcggcagc 541 acatacagcg attttaaatg gcattggtac cattttgacg gaaccgattg ggacgagtcc 601 cgaaagctga accgcatcta taagtttcaa ggaaaggctt gggattggga agtttccaat 661 gaaaacggca actatgatta tttgatgtat gccgacatcg attatgacca tcctgatgtc 721 gcagcagaaa ttaagagatg gggcacttgg tatgccaatg aactgcaatt ggacggtttc 781 cgtcttgatg ctgtcaaaca cattaaattt tcttttttgc gggattgggt taatcatgtc 841 agggaaaaaa cggggaagga aatgtttacg gtagctgaat attggcagaa tgacttgggc 901 gcgctggaaa actatttgaa caaaacaaat tttaatcatt cagtgtttga cgtgccgctt 961 cattatcagt tccatgctgc atcgacacag ggaggcggct atgatatgag gaaattgctg 1021 aacggtacgg tcgtttccaa gcatccgttg aaatcggtta catttgtcga taaccatgat 1081 acacagccgg ggcaatcgct tgagtcgact gtccaaacat ggtttaagcc gcttgcttac 1141 gcttttattc tcacaaggga atctggatac cctcaggttt tctacgggga tatgtacggg 1201 acgaaaggag actcccagcg cgaaattcct gccttgaaac acaaaattga accgatctta 1261 aaagcgagaa aacagtatgc gtacggagca cagcatgatt atttcgacca ccatgacatt 1321 gtcggctgga caagggaagg cgacagctcg gttgcaaatt caggtttggc ggcattaata 1381 acagacggac ccggtggggc aaagcgaatg tatgtcggcc ggcaaaacgc cggtgagaca 1441 tggcatgaca ttaccggaaa ccgttcggag ccggttgtca tcaattcgga aggctgggga 1501 gagtttcacg taaacggcgg gtcggtttca atttatgttc aaagatag Explain... a). How will you construct the forward and reverse primers (do not forget 3 Nucleotide Sequence before the Restriction Enzyme part of the primer)? Explain why and how you made the primers like you did.

Utilizing Hind III and EcoR V Restriction Enzyme with Pet41 and the following gene of interest... a tgaaacaaca aaaacggctt tacgcccgat tgctgacgct gttatttgcg 61 ctcatcttct tgctgcctca ttctgcagca gcggcggcaa atcttaatgg gacgctgatg 121 cagtattttg aatggtacat gcccaatgac ggccaacatt ggaagcgttt gcaaaacgac 181 tcggcatatt tggctgaaca cggtattact gccgtctgga ttcccccggc atataaggga 241 acgagccaag cggatgtggg ctacggtgct tacgaccttt atgatttagg ggagtttcat 301 caaaaaggga cggttcggac aaagtacggc acaaaaggag agctgcaatc tgcgatcaaa 361 agtcttcatt cccgcgacat taacgtttac ggggatgtgg tcatcaacca caaaggcggc 421 gctgatgcga ccgaagatgt aaccgcggtt gaagtcgatc ccgctgaccg caaccgcgta 481 atttcaggag aacacctaat taaagcctgg acacattttc attttccggg gcgcggcagc 541 acatacagcg attttaaatg gcattggtac cattttgacg gaaccgattg ggacgagtcc 601 cgaaagctga accgcatcta taagtttcaa ggaaaggctt gggattggga agtttccaat 661 gaaaacggca actatgatta tttgatgtat gccgacatcg attatgacca tcctgatgtc 721 gcagcagaaa ttaagagatg gggcacttgg tatgccaatg aactgcaatt ggacggtttc 781 cgtcttgatg ctgtcaaaca cattaaattt tcttttttgc gggattgggt taatcatgtc 841 agggaaaaaa cggggaagga aatgtttacg gtagctgaat attggcagaa tgacttgggc 901 gcgctggaaa actatttgaa caaaacaaat tttaatcatt cagtgtttga cgtgccgctt 961 cattatcagt tccatgctgc atcgacacag ggaggcggct atgatatgag gaaattgctg 1021 aacggtacgg tcgtttccaa gcatccgttg aaatcggtta catttgtcga taaccatgat 1081 acacagccgg ggcaatcgct tgagtcgact gtccaaacat ggtttaagcc gcttgcttac 1141 gcttttattc tcacaaggga atctggatac cctcaggttt tctacgggga tatgtacggg 1201 acgaaaggag actcccagcg cgaaattcct gccttgaaac acaaaattga accgatctta 1261 aaagcgagaa aacagtatgc gtacggagca cagcatgatt atttcgacca ccatgacatt 1321 gtcggctgga caagggaagg cgacagctcg gttgcaaatt caggtttggc ggcattaata 1381 acagacggac ccggtggggc aaagcgaatg tatgtcggcc ggcaaaacgc cggtgagaca 1441 tggcatgaca ttaccggaaa ccgttcggag ccggttgtca tcaattcgga aggctgggga 1501 gagtttcacg taaacggcgg gtcggtttca atttatgttc aaagatag Explain... a). How will you construct the forward and reverse primers (do not forget 3 Nucleotide Sequence before the Restriction Enzyme part of the primer)? Explain why and how you made the primers like you did.

Human Anatomy & Physiology (11th Edition)

11th Edition

ISBN:9780134580999

Author:Elaine N. Marieb, Katja N. Hoehn

Publisher:Elaine N. Marieb, Katja N. Hoehn

Chapter1: The Human Body: An Orientation

Section: Chapter Questions

Problem 1RQ: The correct sequence of levels forming the structural hierarchy is A. (a) organ, organ system,...

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Utilizing Hind III and EcoR V Restriction Enzyme with Pet41 and the following gene of interest...

a tgaaacaaca aaaacggctt tacgcccgat tgctgacgct gttatttgcg 61 ctcatcttct tgctgcctca ttctgcagca gcggcggcaa atcttaatgg gacgctgatg 121 cagtattttg aatggtacat gcccaatgac ggccaacatt ggaagcgttt gcaaaacgac 181 tcggcatatt tggctgaaca cggtattact gccgtctgga ttcccccggc atataaggga 241 acgagccaag cggatgtggg ctacggtgct tacgaccttt atgatttagg ggagtttcat 301 caaaaaggga cggttcggac aaagtacggc acaaaaggag agctgcaatc tgcgatcaaa 361 agtcttcatt cccgcgacat taacgtttac ggggatgtgg tcatcaacca caaaggcggc 421 gctgatgcga ccgaagatgt aaccgcggtt gaagtcgatc ccgctgaccg caaccgcgta 481 atttcaggag aacacctaat taaagcctgg acacattttc attttccggg gcgcggcagc 541 acatacagcg attttaaatg gcattggtac cattttgacg gaaccgattg ggacgagtcc 601 cgaaagctga accgcatcta taagtttcaa ggaaaggctt gggattggga agtttccaat 661 gaaaacggca actatgatta tttgatgtat gccgacatcg attatgacca tcctgatgtc 721 gcagcagaaa ttaagagatg gggcacttgg tatgccaatg aactgcaatt ggacggtttc 781 cgtcttgatg ctgtcaaaca cattaaattt tcttttttgc gggattgggt taatcatgtc 841 agggaaaaaa cggggaagga aatgtttacg gtagctgaat attggcagaa tgacttgggc 901 gcgctggaaa actatttgaa caaaacaaat tttaatcatt cagtgtttga cgtgccgctt 961 cattatcagt tccatgctgc atcgacacag ggaggcggct atgatatgag gaaattgctg 1021 aacggtacgg tcgtttccaa gcatccgttg aaatcggtta catttgtcga taaccatgat 1081 acacagccgg ggcaatcgct tgagtcgact gtccaaacat ggtttaagcc gcttgcttac 1141 gcttttattc tcacaaggga atctggatac cctcaggttt tctacgggga tatgtacggg 1201 acgaaaggag actcccagcg cgaaattcct gccttgaaac acaaaattga accgatctta 1261 aaagcgagaa aacagtatgc gtacggagca cagcatgatt atttcgacca ccatgacatt 1321 gtcggctgga caagggaagg cgacagctcg gttgcaaatt caggtttggc ggcattaata 1381 acagacggac ccggtggggc aaagcgaatg tatgtcggcc ggcaaaacgc cggtgagaca 1441 tggcatgaca ttaccggaaa ccgttcggag ccggttgtca tcaattcgga aggctgggga 1501 gagtttcacg taaacggcgg gtcggtttca atttatgttc aaagatag

Explain...

a). How will you construct the forward and reverse primers (do not forget 3 Nucleotide Sequence before the Restriction Enzyme part of the primer)? Explain why and how you made the primers like you did.

b). Which Pet41 (a, b or c) would you use for cloning the desired gene? How would you get the gene in the right reading frame

T7 promoter
lac operator
GST Tag
Munl
AATTAATACGACTCACTATAGGGGAATTGTGAGCGGATAACAATTCCCCTCTAGAAATAATTTTGTTTAACTTTAAGAAGGAGATATACATATGT CCCCT
Met Ser Pro
Xbal
Avril
Spel
S.Tag
rbs
His-Tag
ATACTAGGTTAT...627bp... GACCATCCTCCAAAATCGGATGGTTCAACTAGTGGTTCTGGTCATCACCATCACCATCACTCCGCGGGTCTGGTGCCACGCGGTAGT
Ile Leu Gly Tyr...209aa. . . Asp His Pro Pro Lys Ser Asp Gly Ser Thr Ser Gly Ser Gly His His His His His His Ser Ala Gly Leu Val Pro Arg Gly Ser
thrombin
PinAl
Bgl || Kpnl
Ndel
Sac Il
S Tag primer #69945-3,
ACTGCAATTGGTATGAAAGAAACCGCTGCTGCTAAATTCGAACGCCAGCACATGGACAGCCCAGATCTGGGTACCGGTGGTGGCTCCGGTGATGACGACGACAAG
Thr Ala Ile Gly Met Lys Glu Thr Ala Ala Ala Lys Phe Glu Arg Gln His Met Asp Ser Pro Asp Leu Gly Thr Gly Gly Gly Ser Gly Asp Asp Asp Asp Lys
enterokinase
Pstl
Eagl
Ncol EcoR V BamHI EcoRI BsrGI Stul Ascl Sse83871 Sacl Sall Hind III Notl
AGTCCCATGGGATATCGGGGATCCGAATTC TGTACAGGCCTTGGCGCGCCTGCAGGCGAGCTCCGTCGACAAGCTTGCGGCCGCACTCGAGCACCACCACCACCACCACCACCACTAATTGATTAA PET-41a(+)
Ser Pro Met Gly Tyr Arg Gly Ser Glu Phe Cys Thr Gly Leu Gly Ala Pro Ala Gly Glu Leu Arg Arg Gln Ala Cys Gly Arg Thr Arg Ala Pro Pro Pro Pro Pro Pro Pro Leu Ile Asp End
Xhol
PshAl
AGTCCCATGGATATCGGGGATCCGAATTC TGTACAGGCCTTGGCGCGCCTGCAGGCGAGCTCCGTCGACAAGCTTGCGGCCGCACTCGAGCACCACCACCACCACCACCACCACTAATTGATTAA PET-41b(+)
Ser Pro Met Asp Ile Gly Asp Pro Asn Ser Val Gln Ala Leu Ala Arg Leu Gln Ala Ser Ser Val Asp Lys Leu Ala Ala Ala Leu Glu His His His His His His His His End
AGTCCCATGGCGATATCGGGGATCCGAATTCTGTACAGGCCTTGGCGCGCCTGCAGGCGAGCTCCGTCGACAAGCTTGCGGCCGCACTCGAGCACCACCACCACCACCACCACCACTAATTGATTAA PET-41c(+)
Ser Pro Met Ala Ile Ser Gly Ile Arg Ile Leu Tyr Arg Pro Trp Arg Ala Cys Arg Arg Ala Pro Ser Thr Ser Leu Arg Pro His Ser Ser Thr Thr Thr Thr Thr Thr Thr Thr Asn End
T7 terminator
His•Tag
Bpu1102 I
TACCTAGGCTGCTAAACAAAGCCCGAAAGGAAGCTGAGTTGGCTGCTGCCACCGCTGAGCAATAACTAGCATAACCCCTTGGGGCCTCTAAACGGGTCTTGAGGGGTTTTTTG
T7 terminator primer #69337-3

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