Chapter 7 Microbial Genetics Plasmids Circular genetic elements that reproduce autonomously and have an extra chromosomal existence 1-1000 KB in size Most are circular double- stranded dna. some linear ds DNA Transmitted from cell to cell via conjugation process Some can integrated into chromosome Can carry a variety of genes for production of toxin, resistance to antibiotics and heavy metals
Chapter 7 Microbial Genetics Plasmids • Circular genetic elements that reproduce autonomously and have an extrachromosomal existence. – 1-1000 KB in size – Most are circular doublestranded DNA, some linear ds DNA – Transmitted from cell to cell via conjugation process – Some can integrated into chromosome – Can carry a variety of genes for production of toxin, resistance to antibiotics and heavy metals
Mutations and Mutants Phenotype: the observable characteristics of an organism designated by a capital letter followed by two small letters, with either a plus and minus superscript to indicate the presence or absence of that property His. Glut et al Genotype: The precise genetic make-up of an organism designated by three lower case letters followed by a capital letter(all in italics) indicating the particular gene involved: hisC, mutations in the hisc gene would be designated as hisEl, hisC2 et al
Mutations and Mutants • Phenotype:the observable characteristics of an organism – designated by a capital letter followed by two small letters, with either a plus and minus superscript to indicate the presence or absence of that property: His- , Glu+ et al • Genotype: The precise genetic make-up of an organism – designated by three lower case letters followed by a capital letter (all in italics) indicating the particular gene involved: hisC, mutations in the hisC gene would be designated as hisC1, hisC2 et al
Isolation of mutants Pigmented and nonpigmented mutants Antibiotic-resistant mutants within the inhibition zone · Genetic marker, such as the expression ofβ- galactosidase that produces the blue color
Isolation of Mutants • Pigmented and nonpigmented mutants • Antibiotic-resistant mutants within the inhibition zone • Genetic marker, such as the expression of bgalactosidase that produces the blue color
Replica plating Technique: Isolation of Mutants Velveteen(锦绒布) ∵ Auxotroph(营养缺陷型) Mutants do not grow Prototroph(原养型) 合-合 growth on complete medium Press plate onto velveteen Velveteen with imprint of all Animal medium Complete medium colonies Velveteen Plastic hoop ∵: Wooden block Transfer imprint of colonies to
Replica Plating Technique: Isolation of Mutants Velveteen (锦绒布） Auxotroph (营养缺陷型) Prototroph (原养型)
Replica plating Technique: Isolation of Mutants FIGURE 7.2 (a) Replica plating method for detection of nutritional mu- tants. (b) Nutritional mutants, as re- vealed by the replica plating method The left photo shows the master plate The colonies not appearing on the epoca plate are marked with an x. The replica plate lacked one nutrient leucine) present in the master plate Penicillin-Selection method Penicillin kills only growing cells. If penicillin is added to a population growing in a medium lacking the growth factor required by the designed mutant, the parent cells will be killed whereas the non-growing mutant cells will be unaffected
Replica Plating Technique: Isolation of Mutants • Penicillin-Selection Method – Penicillin kills only growing cells. If penicillin is added to a population growing in a medium lacking the growth factor required by the designed mutant, the parent cells will be killed, whereas the non-growing mutant cells will be unaffected
Mutants that are detectable Non-motile Non-capsulated Rough colony Nutritional Sugar fermentation Drug resistant Ⅴ irus resistant Temperature sensitive gmentiess Cold sensitive
Mutants that are detectable • Non-motile • Non-capsulated • Rough colony • Nutritional • Sugar fermentation • Drug resistant • Virus resistant • Temperature sensitive • Pigmentless • Cold sensitive
5·TAc… Molecular ATG…5 MUTATION Normal ba f asis o Mutation A c T A C T G A T A T G · Point mutation ↓↓↓↓ Transcription Missense mutation A C U A G U AU U A C Nonsense mutation Asparagine 天so op Tyrosine Tyrosine codon 冬酰胺 codon 酪氨酸 Silent mutation Translation Faulty Incomplete Normal Normal Protein Protein Protein Protein Missense Nonsense Silent Wild type mutation mutation FIGURE 7.3 Possible effects of base pair substitution in a gene cod ing for a protein: three different protein products from changes in the DNA for a single codon
Molecular basis of Mutation • Point Mutation – Missense mutation – Nonsense mutation – Silent mutation 天冬酰胺 酪氨酸
Frameshift mutations GIC C G AT Insertion mutation TA CGG ACIC AA T AC T Altered reading frame 个m Normal DNA T GC CITIG T TA TG T A CG GAC A AT AC Normal reading frame Deletion Deletion mutation T GIC C GIT T AIT Fi T A CIG G CIA A TA Altered reading frame by Occur in protein-encoding gene that including the reading frame If occurs in promoter region, it is not frameshift
Frameshift mutations • Occur in protein-encoding gene that including the reading frame • If occurs in promoter region, it is not frameshift
Back mutations or reversions Point mutations are reversible Revertant wild type phenotype that was lost in the mutant is restored Same site revertants True revertants Second site mutation Suppressor mutation: restore the wild type phenotype by mutation somewhere else
Back Mutations or Reversions • Point Mutations are reversible • Revertant – wild type phenotype that was lost in the mutant is restored. • Same site revertants • True revertants • Second site mutation – Suppressor mutation: restore the wild type phenotype by mutation somewhere else
Mutations involving many base pairs Deletions: mutations in which a region of the dna has been eliminated Insertions occur when new bases are added to the dNA, can be a sing de base or many bases(called insertion sequences) Translocation: large section of chromosomal dna is moved to a new location
Mutations involving many base pairs • Deletions: mutations in which a region of the DNA has been eliminated • Insertions: occur when new bases are added to the DNA, can be a single base or many bases (called insertion sequences) • Translocation: large section of chromosomal DNA is moved to a new location