Which of the restriction enzymes listed in the table below produces blunt-end fragments? Enzyme Cleavage site Alul AG|CT Hindll AJAGCTT BamHI G|GATCC EcoRI GJAATTC BgllI A|GATCA Alul O BamHI O Bell O EcoRI O Hindll
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- What restriction enzyme (or enzymes) would you use to cut the following... (Gene of Interest is Bolded) 1 tctagagtca 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…#1 HindII --- 5’ GTC ↓ GAC 3’ 5’ ACGACGTAGTCGACTTATTAT GTCGACCCGCCGCGTGTCGACCATCA 3’ 3’ TGCTGCATCAGCTGAATAATACAGCTGGGCGGCGCACAGCTGGTAGT 5’ Restriction enzyme: Recognition sequence: Number of pieces of DNA: Type of cut:#4 BamI --- 5’ CCTAG ↓G 3’ 5’ ACGCCTAGGACGTATTATCCTAGGTAT CCGCCGCCGT CATCA 3’ 3’ TGCGGATCCTGCATAATAGGATCCATAGGCGGCGGCAGTAGT 5’ Restriction enzyme: Recognition sequence: Number of pieces of DNA: Type of cut:
- #2 EcoRI --- 5’ G ↓AATTC 3’ 5’ ACG ACGTATTAGAATTCTTA TCCGCCGCCGGAATTCT CATCA 3’ 3’ TGC TGCATAATCTTAAGAATAGGCGGCGGCCTTAAGAGTAGT 5’ Restriction enzyme: Recognition sequence: Number of pieces of DNA: Type of cut:#3 HaelII --- 5’ CC ↓ GG 3’ 5’ ACGCCGGCCGTATTAT CCGGATCCGCCG CCGGCTGTCCCGGATCA 3’ 3’ TGCGGCCGGCATAATAGGCCTAGGCGGCGGCCGACAGGGCCTAGT 5’ Restriction enzyme: Recognition sequence: Number of pieces of DNA: Type of cut:18. You want to express hemoglobin beta (HBB) in a bacteria model using a vector. You would like to amplify the following gene by PCR. GTAGAATATG ATAAGCGAAA CTGCAAATCG CGTTTGGGGC GATACAAGTA GTGTACGCGG ACCGCGCCGA GCGGGCTATG GTTTCTCCAC TATCAGTTCT TCTCCTGTCC CAGCGAAGTC GAGGTCCAGC CCTACGGTAC ATAACTAACA CGGTTTGAAG AAGATACGAT CTTACGAAGT AAAGAAAATT TGTAGTCAGC CCGGTTCGCT TGTGTCCAGC TAATCGATTG ATTGGCCCCA GCAGGCGAGA TGAACATAGT CATGCGCTGT CTAATAGCCC ATTTGACGTG TAGGTGGCGC TTTTATTTCT GAGGTGGAAA T a) If the primers were both 20 nucleotides long, what are the sequences for both PCR primers that will amplify only the highlighted region? Be sure to label the 5' and 3' ends. (4 points) b) What is the length (in bp) of the entire region amplified by their primers (i.e. the amplicon length)? (Note: the spacing in the above sequence is intentionally uniform) (2 points) c) If you start with 300 copies template DNA how many copies would you expect to produce if you ran the PCR for 22 cycles? Show your work.…
- FIGURE 2 shows the only suitable DNA restriction site in a plasmid DNA vector that can be cleaved. 5'.GTAATCCGGATCCGAAATCCCCCGG... 3' 3.CATTAGGCCTAGGCTTTAGGGGGCCC... 5 FIGURE 2 Identify the most suitable enzymeto be used as a restriction enzyme forthis purpose. O Ligase O Smal O EcoRI О ЕсoRVKha Vu Danels Include: 8DX : Safehy Jor bromie tnto lab repart! Name Section date sheet MAPPING PRACTICE #1 Below is a restriction map for the plasmid PGEN 101 (total length = 20 Kb). Using this map as a guide, give the number of restriction fraqments along with their associated lengths that would result from digesting PGEN 101 with the restriction enzymes EcoRI, BamHI and a combination of ECORI and BamHI. BamHI 3.2 Kb 1.7 Kb EcoRI BamHI PGEN 101 8.7 Kb 5.5 Kb .9 Kb EcoRI ECORI DIGESTION PERFORMED SIZES OF FRAGMENTS OBTAINED 10.4 kb , 0.9kb, 8.7 Kb EcoRI 3.2 Kb, 16. 8kb BamHI EcoRI + BamHIUtilizing 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…
- Given below is the DNA template. What are the gene products? 3’ TACCGGCCTATCTAGGGCCATGGCTTAATTCCC 5’ 5’ ATGGCCGGATAGATCCCGGTACCGAATTAAGGG3’Please determine the relative location of the three restriction enzymes. Enzyme Number of fragments Size (kb) Xbal 2 17, 21 Xhol 2. 5, 33 Xbal/Xhol 5, 12, 21 Kpnl 6, 7, 58 Xbal/Kpnl 6, 7, 8, 17restriction recognition site for: (you can also find this on the internet). Be sure to give Using the chart posted to the left of the microwave in lab, what is the 3. restriction recognition site for: (you can also find this on the internet). Be sure te the sequence and indicate the cut sites with an arrow. EcoR1: Hindlll: BamH1: