Biotechnology and its Applications Set-4

Q1. In a PCR reaction starting with $N_0=100$ template molecules and assuming ideal amplification (each cycle doubles the number of molecules), what is the minimum number of cycles $n$ required to obtain at least $1\times10^6$ copies? (Use $N=N_0 2^n$.)

Q2. A recombinant human glycoprotein requires N‑linked glycosylation and correct disulfide bond formation to be biologically active. Which of the following expression systems is most likely to produce a biologically active, human‑like glycosylated form with proper post‑translational modifications?

Q3. In a paternity test three STR loci were analyzed. The allele sizes (bp) observed are:
Locus I — Mother: $110,130$; Child: $130,140$; Man A: $110,150$; Man B: $140,150$.
Locus II — Mother: $200,220$; Child: $220,210$; Man A: $200,210$; Man B: $210,230$.
Locus III — Mother: $310,330$; Child: $310,340$; Man A: $340,350$; Man B: $300,340$.
Based on Mendelian inheritance at these loci, which man is the biological father of the child?

Q4. Assertion (A): Expression of human therapeutic glycoproteins in Saccharomyces cerevisiae yields glycosylation patterns identical to those produced in human cells.
Reason (R): Yeast N‑linked glycosylation predominantly adds high‑mannose oligosaccharide chains and lacks terminal sialic acids, making them structurally different from human N‑glycans.
Choose the correct option regarding A and R.

Q5. In a chemostat the microbial growth follows Monod kinetics: $\mu=\mu_{\max}\dfrac{S}{K_s+S}$. For a culture with $\mu_{\max}=0.6\ \mathrm{h^{-1}}$, $K_s=0.2\ \mathrm{g\,L^{-1}}$, substrate in feed $S_0=10\ \mathrm{g\,L^{-1}}$, yield coefficient $Y=0.5\ \mathrm{g\ biomass/g\ substrate}$ and dilution rate $D=0.4\ \mathrm{h^{-1}}$, steady state satisfies $D=\mu$ and biomass $X=Y(S_0-S)$. What is the steady‑state biomass concentration $X$ (g L^{-1})?

Q6. A circular plasmid of size $8\ \text{kb}$ was analysed by restriction digestion. Digestion with enzyme R1 alone produced two fragments of $3\ \text{kb}$ and $5\ \text{kb}$. Digestion with enzyme R2 alone produced two fragments of $4\ \text{kb}$ and $4\ \text{kb}$. A double digest with R1 and R2 produced three fragments of $1\ \text{kb}$, $3\ \text{kb}$ and $4\ \text{kb}$. Which of the following arrangements of restriction sites on the plasmid best explains these results?

Q7. In a PCR reaction with perfect efficiency ($100\%$) each cycle doubles the number of target DNA molecules. If the reaction starts with $10^3$ copies of template and runs for $20$ cycles, the final expected copy number is $N = N_0\times 2^{n}$. What is the numerical value of $N$ after $20$ cycles?

Q8. A transgenic plant carries a single-copy insertion of a $1\ \text{kb}$ transgene at a defined locus. Primers flanking the insertion site amplify a $0.5\ \text{kb}$ product from the wild-type allele and a $1.5\ \text{kb}$ product from the allele containing the insertion. An F1 heterozygote (one wild-type allele, one insertion allele) is selfed to produce an F2 population. What fraction of F2 plants are expected to show only the $1.5\ \text{kb}$ band (i.e., be homozygous for the insertion)?

Q9. In a paternity test three STR loci (A, B, C) were scored (allele sizes in base pairs). The observed alleles are:

• Locus A: Mother (M) = $120,140$; Child (C) = $120,160$; Alleged father F1 = $150,160$; Alleged father F2 = $130,150$.
• Locus B: M = $200,220$; C = $200,240$; F1 = $210,240$; F2 = $220,240$.
• Locus C: M = $300,340$; C = $340,360$; F1 = $360,380$; F2 = $350,360$.
  Based on Mendelian inheritance (child receives one allele from mother and one from father at each locus), which individual is most consistent with being the biological father?

Q10. A researcher uses viral transduction and models the distribution of viral particles per cell with a Poisson process. For multiplicity of infection (MOI) $m$, the probability that a cell receives zero viral particles is $P(0)=e^{-m}$. The researcher needs at least $95\%$ of cells to be infected (i.e., receive one or more particles). What minimum MOI should be used to achieve this target? (Give the approximate value derived from $1-e^{-m}\ge 0.95$.)

Q11. In an ideal PCR reaction where each cycle doubles the number of DNA copies, starting with $50$ template molecules, how many copies will be present after $20$ cycles? Use $N = N_0 \times 2^n$.

Q12. A circular plasmid of total size $11\ \text{kb}$ is analyzed by restriction digestion. Single digest with enzyme X yields fragments of $3\ \text{kb}$ and $8\ \text{kb}$. Single digest with enzyme Y yields fragments of $4\ \text{kb}$ and $7\ \text{kb}$. Double digest with X and Y yields fragments of $1\ \text{kb}$, $2\ \text{kb}$, $3\ \text{kb}$ and $5\ \text{kb}$. Which of the following gives the correct order of restriction sites around the plasmid (clockwise)?

Q13. In a bacterial transformation, $0.01\ \mu\text{g}$ of plasmid DNA was used. After recovery the total volume was $1.0\ \text{mL}$; $0.1\ \text{mL}$ of this was plated and produced $120$ colonies. Using $TE = \dfrac{\text{total transformants}}{\text{DNA amount }(\mu\text{g})}$, what is the transformation efficiency in transformants per microgram?

Q14. A T0 transgenic plant carrying a dominant selectable marker was selfed. Among 156 T1 seedlings, 145 were resistant and 11 were sensitive. Which of the following is the most likely explanation for this segregation?

Q15. Two independent transgenic lines (A and B) carry the same transgene under the same promoter. Southern blot shows line A has a single-copy insertion and high mRNA and protein; line B has multiple tandem copies but negligible mRNA and no protein. Bisulfite sequencing of line B reveals heavy methylation of the transgene promoter region. Which mechanism best explains loss of expression in line B?