marks 764-780 763 ence 366-692 ence 327-344 249-293 ence 158-217 140-157 26-72 1171-2250 3684 4445-5302 5434-5889 b(+) a

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Marks 764 780 763 Ence 366 692 Ence 327 344 249 293 Ence 158 217 140 157 26 72 1171 2250 3684 4445 5302 5434 5889 B A 1 (57.51 KiB) Viewed 31 times

Marks 764 780 763 Ence 366 692 Ence 327 344 249 293 Ence 158 217 140 157 26 72 1171 2250 3684 4445 5302 5434 5889 B A 2 (16.69 KiB) Viewed 31 times

marks 764-780 763 ence 366-692 ence 327-344 249-293 ence 158-217 140-157 26-72 1171-2250 3684 4445-5302 5434-5889 b(+) and PET-32c(+) 32a(+) (shown) with ons: pET-32b(+) is a act 1bp from each site 8. PET-32c(+) is a Ibp to each site 8 except for EcoR V. T7 promoter Sca (4995) Pvulands) Pst (4760) Bsa l(4576) Eam1105 (4357) AlwN (4038) Dra Ill(5658) 4445-5302) Msc1 BspLU11 (3622) Sap (3506) Bst1107 1(3393) TH191 19967). BspG (3148) Xbal lac operator TAATACGACTCACTATAGGGGAATTGTGAGCGGATAACAATTCCCCTCTAGAAATA On (3684) 11 origin (5434-5889) Bou11021 Bpu1102 (80) PET-32a(+) (5900bp) TAACTTTAAGAAGGAGA GGTATGAAAGAAACCGCTGCTGCTAAATTCGAACGCCAGCACATGGACAGCCCAGATETOGGTACCOACGACGACGACAAG GlyMetLysGluThrAlaAloklaysPheGluArgüinHistetAspSerProAspleuGlyThrAspAspAspAspLys rbs LysProbluargLysLeuser TrpLeuleuProProLeuserAnAnEnd His-Tag AloMetAlolleSer AspProAsnSer Ser SerValAsplysLeuAloAlokToLeuttuHisHisHisHiHiHind His-Tag Trx-Tag TATACATATGAGE 315bp CTGGCCGGTTCTGOTTCTGOCCATATGCACCATCATCATCATCATTCTTCTGGTCTGGTGCCACGCGCTICT MetSer 10500 LeuAloGlySerGlySerGlyMetHisKisHiSHISHISHISSerSerGlyLeuVolProArgGlySer S-Tag primer #69945-3 Ball Kon S-Tag Nap V Bgl (241) -Kpn 1(238) -Nsp V(268) Msc (351) Rsr Il(589) 1XA (366-692) T7 terminator primer #69337-3 PET-32a-c(+) cloning/expression region Psp5 (228) Ava (158) Xho (158) Eag (166) Not (166) Hind (173) Sal (179) Sac (190) EcoR (192) BamH (198) EcoR V(206) Nco (212) thrombin PET-32a(+) Eagl Aval Nool EcoR V BamHI EcoRI Sacl Sall Hind Ill Not I Xhol GCCATOGETGATATCGGATCEGAATICOAGCTCCGTCGACAAGCTTGCOGECGCACTCGAGCACCACCACCACCACCACTGAGATCCOGCTGCTA AloMetAloAsplieGlySerGluPheGluLeuArgArgGInA laCysGlyArgThe ArgAloProProProProfraLeuArgSerGlycysEnd) GCCATGGEGATATCGEATCEGAATTCGAGCTCCGTCGACAAGETTGCGGCCGCACTCGASCACCACCACCACCACCACTGAGATCCOGETOCTAA enterokinase Xba I(729) CAAAGCCCGAAAGGAAGETGAGTTGGETOCTOCCACCGETGAGSCAATAACTAGCATAACCCCTTGGGGCCTCTAAACGGOTCTTGAGGGGTTTTTT SgrA (840) PshA (2366) lacl (1171-2250) Sph 1996) EcoN (1056) ApaB (1205) Mlu (1521) Bel (1535) GCCATOGGATATCTGTGGATCCGAATTEGAGCTCCGTCGACAAGCTTOCGGCCGCACTCGASCACCACCACCACCACCACTGAGATCCOGCTGCTAA PET-32c(+) AloMetGlyTyrLeutrolleArelleArgAloProSer ThrSerLeuAroPratisSer SerThrThrTheTheThe ThrGiul leAreleuleuTh T7 terminator BstE (1702) Bmg (1730) Apa (1732) BssH (1932) Hpa (2027) PET-32b(+)
4.1. How many fragments will form if pET32a(+) is digested simultaneously with Sacl and Bcll. [1] 4.2. Write down the base pairs that result at the 5' and 3' ends of the shortest fragment [3] 4.3. Restriction enzymes often recognize palindromic sequences. What is a palindromic sequence? [2] 4.4. Give three restriction enzymes found on the pET32(a+) multiple cloning site. [3]